Localized molecular and ionic transport to and from tissues

ABSTRACT

The present invention relates to methods and devices used for the formation of microconduits in a tissue. The term “microconduit” refers to a small opening, channel, or hole into, or through, a tissue, that allows transfer of materials by liquid flow, and by electrophoresis, the microconduit being formed upon impact of a plurality of accelerated microparticles with the surface of the tissue. A method is described for forming at least one microconduit in tissue including the steps of: accelerating a plurality of microparticles to a velocity that causes the microparticles to penetrate a region of tissue surface upon impingement of the microparticles on the tissue surface; and directing the microparticle towards the region of tissue surface, thereby causing the microparticles to penetrate the tissue and form a microconduit in the tissue. According to an embodiment, microparticles are accelerated by being hit with a moving, solid surface. In another embodiment, microparticles are accelerated by a flowing gas or liquid. Also described are methods and devices for using microconduits to deliver therapeutic molecules and ions into tissue, or for extraction of chemical analytes out of tissue. Also described is a method of nail piercing to accommodate jewelry.

RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. ProvisionalApplication No. 60/209,985, filed on Jun. 8, 2000, the teachings ofwhich are incorporated herein by reference in their entirety.

BACKGROUND OF THE INVENTION

[0002] Transdermal drug delivery, as the term is used generally, refersto permeation of the stratum corneum, the tough outer barrier of theskin, by a pharmaceutically active molecule. The stratum corneum, thethin (approximately 20 μm) outer layer of the epidermis, is dead tissuecontaining both multilamellar lipid barriers, and tough protein-basedstructures.

[0003] The epidermis, directly beneath the stratum corneum, also behavesas a lipid barrier. The dermis, directly beneath the epidermis, ispermeable to many types of solutes. In the administration of a drug bytopical application to skin, lipid-soluble drug molecules dissolve intoand diffuse through the skin's multilamellar lipid bilayer membranesalong a concentration gradient by virtue of the drug molecules'solubility in the lipid bilayer. Transdermal drug delivery may betargeted to a tissue directly beneath the skin, or to capillaries forsystemic distribution within the body by the circulation of blood.

[0004] The term “transdermal drug delivery” usually excludes hypodermicinjection, long term needle placement for infusion pumps, and otherneedles which penetrate the skin's stratum corneum. Thus, transdermaldrug delivery is generally regarded as minimally invasive. However, thelow rate of transport of therapeutic molecules through the stratumcorneum remains a common clinical problem.

[0005] Transdermal delivery of only a limited number of lipophilic drugsis commercially available. Existing methods include, for example, theuse of wearable “patches,” a passive transdermal drug delivery methodthat tends to be slow, and difficult to control.

[0006] Another method includes the use of a “gene gun,” to accelerate 20to 70 μm diameter drug particles, or smaller DNA-coated gold particles,to supersonic velocities, such that the particles pass through thestratum corneum into the epidermis or dermis. A single particle, 20 μmto 70 μm, in diameter, such as used in the gene gun, when fired at thestratum corneum at supersonic speeds, ruptures and tears through thetissues of the stratum corneum, epidermis and dermis, stopping andremaining at some depth which is determined by the initial velocity andmass of the particle. The resulting path through the above-mentionedtissues may be in the range of 1 μm to perhaps 30 μm because the tissuesare elastic to various degrees, depending on the individual. Thesemi-static analogue is to pierce a rubber sheet with a common pin, 750μm in diameter. When pulled out of the rubber sheet, the resultantopening size is less than 1 μm, or perhaps not open at all. This isbecause the pin has torn the rubber sheet and pushed it aside, due tothe rubber sheet's elasticity (ability to get out of the way), as thepin is forced through. As in the analogue, because of the elasticity ofskin, use of the gene gun does not form microconduits in the skinbecause the tissue is only temporarily pushed aside as a particle isforced through the skin.

[0007] Examples of transdermal drug delivery methods presently beinginvestigated include the use of ultrasound (sonophoresis) to causecavitation in the stratum corneum; laser ablation of a small region ofthe stratum corneum, thereby providing access to the epidermis; the useof microneedles to create openings in the stratum corneum; the use ofelectrical methods, including low voltage iontophoresis, whereintransport is believed to occur through pre-existing aqueous pathways;and the use of high voltage pulses to cause electroporation of the skin.There are disadvantages associated with each of these methods. Forexample, often the rate of transport of molecules tends to diminishrapidly with increasing molecular size. Other disadvantages include painand discomfort, skin irritation, the high cost and the large size ofequipment required, and the potential for breaking off needles, whichmight remain imbedded in the skin.

[0008] Also, a common problem encountered in using establishedtechniques such as subcutaneous and intradermal injection to delivervaccines, is the inaccurate placement of the immunizing material withrespect to the epidermal and dermal antigen-presenting cells, or withrespect to keratinocytes. There is also a long-standing need for aneffective method to deliver therapeutic agents to treat a fungalinfection of the tissue underlying nail tissue of fingers and toes.

[0009] An existing problem with currently used methods of makingbiopotential measurements and other electrical measurements at thesurface of the skin of a living organism is that the measurements areoften degraded by motion and by other potentials that are associatedwith the skin. Techniques such as microscission or stripping of thestratum corneum of the skin can significantly improve the quality ofsuch electrical measurements. However, mechanical alteration of the skinis highly undesirable, because it is difficult to control the degree ofalteration; mechanical alteration can cause pain and discomfort, and canlead to infection. Therefore, there is a need for improved methods ofmaking biopotential measurements at the surface of the skin.

[0010] The present invention satisfies these needs by providing, forexample, an improved method of delivery of therapeutic agents to atissue; an improved method of transdermal delivery of therapeuticagents; an improved method for delivering therapeutic agents to tissueunderlying nail tissue; an improved method for obtaining samples ofinterstitial fluid or blood for sensing of analytes within the extractedfluid, including the measurement of analytes while within themicroconduit; and an improved method of making biopotentialmeasurements.

SUMMARY OF THE INVENTION

[0011] The present invention relates to methods and devices for formingmicroconduits in a tissue. The invention, inter alia includes thefollowing, alone or in combination. In one embodiment, a method forforming at least one microconduit in tissue includes the steps of:accelerating a plurality of microparticles to a velocity that causes themicroparticles to penetrate a region of tissue surface upon impingementof the microparticles on the tissue surface; directing themicroparticles towards the region of tissue surface, thereby causing themicroparticles to penetrate the tissue; and scissioning the tissue withthe impinging microparticles, thereby forming a plurality of freemicrotissue particles, and thereby forming a microconduit.

[0012] In another embodiment, a method for forming at least one openingin the stratum corneum of skin includes: accelerating a plurality ofmicroparticles to a velocity that causes the microparticles to penetratea region of the skin surface upon impingement of the microparticles onthe skin surface; directing the microparticles towards the region ofskin surface, thereby causing the microparticles to penetrate the skin;scissioning the skin with the impinging microparticles, thereby forminga plurality of free microtissue particles, and thereby forming amicroconduit.

[0013] The invention also relates to a method of delivery of atherapeutic molecule or ion to tissue, the method including the stepsof: accelerating a plurality of microparticles to a velocity that causesthe microparticles to penetrate a region of tissue surface uponimpingement of the microparticles on the tissue surface; directing themicroparticles towards the region of tissue surface, thereby causing themicroparticles to penetrate the tissue; scissioning the tissue with theimpinging microparticles, thereby forming a plurality of freemicrotissue particles, and thereby forming a microconduit; andadministering at least one therapeutic molecule or ion by directing thetherapeutic molecule or ion into at least one microconduit, therebydelivering a therapeutic molecule or ion to tissue.

[0014] In another embodiment, a method of extracting an analyte from atissue includes: accelerating a plurality of microparticles to avelocity that causes the microparticles to penetrate a region of atissue surface upon impingement of the microparticles on the tissuesurface; directing the microparticles towards the region of tissuesurface, thereby causing the microparticles to penetrate the tissue;scissioning the tissue with the impinging microparticles, therebyforming a plurality of free microtissue particles, and thereby forming amicroconduit; and removing the analyte from the tissue through themicroconduit, thereby extracting the analyte from the tissue.

[0015] The invention also relates to a method for forming a molecularmatrix within at least one microconduit, the method including the stepsof: accelerating a plurality of microparticles to a velocity that causesthe microparticles to penetrate a region of a tissue surface uponimpingement of the microparticles on the tissue surface; directing themicroparticles towards the region of tissue surface, thereby causing themicroparticles to penetrate the tissue; scissioning the tissue with theimpinging microparticles, thereby forming a plurality of freemicrotissue particles, and thereby forming a microconduit; and directinga molecular matrix into the microconduit, thereby forming a molecularmatrix within the microconduit.

[0016] Another embodiment of the invention is a method of transdermaldelivery of a therapeutic molecule or ion, the method including thesteps of: accelerating a plurality of non-drug containing microparticlesto a velocity that causes the microparticles to completely penetrate aregion of a skin surface upon impingement of the microparticles on theskin surface; directing the microparticles towards the region of theskin surface, thereby causing the microparticles to penetrate the skin;scissioning the skin with the impinging microparticles, thereby forminga plurality of free microtissue particles, and thereby forming amicroconduit; and administering at least one therapeutic molecule or ionby directing the therapeutic molecule or ion into at least onemicroconduit, thereby delivering the therapeutic molecule or ion throughthe stratum corneum and into the skin.

[0017] The invention also relates to a method for making one or morebiopotential measurements across the skin, the method including thesteps of accelerating a plurality of microparticles to a velocity thatcauses the microparticles to penetrate a region of a skin surface uponimpingement of the microparticles on the skin surface; directing themicroparticles towards the region of skin surface, thereby causing themicroparticles to penetrate the skin; scissioning the skin with theimpinging microparticles, thereby forming a plurality of freemicrotissue particles, and thereby forming a microconduit; placing atleast two electrodes in electrical connection with the skin with atleast one electrode at the microconduit; and making a biopotentialmeasurement across the skin.

[0018] In one embodiment, the biopotential measurement is anelectrocardiogram. In a particular embodiment, the electrocardiogrammeasurement is obtained during exercise stress testing. In yet anotherembodiment, the biopotential measurement is an electromyogram. Theinvention also relates to the use of microconduits made according to anembodiment for making biopotential measurements suitable forneuromuscular testing. In one embodiment, the biopotential measurementis an electroencephalogram to monitor anaesthesia.

[0019] In a particular embodiment, a method of delivering at least onemolecule to tissue includes the step of storing the molecule in at leastone puncturable capsule in proximity to at least one microconduit. Thestored molecule, according to an embodiment, may be included in apharmaceutically acceptable carrier.

[0020] The invention also relates to a mask for defining at least onelocalized area of a tissue surface region for formation of amicroconduit by microparticle impingement. The mask includes a membranethat has a thickness in a range of between about one micrometer andabout one thousand micrometers; at least one microhole in the membrane,the microhole having a diameter in a range of between about threemicrometers and about one thousand micrometers. The embodiment furtherincludes a means for positioning the membrane against the tissuesurface, on the tissue surface, or near the tissue surface. In aparticular embodiment, the mask is conformable to the tissue surface.

[0021] The invention also relates to a process for forming at least onemicroconduit through nail tissue including the steps of: accelerating aplurality of microparticles to a velocity that causes the microparticlesto penetrate a region of the nail tissue surface upon impingement of themicroparticles on the nail tissue surface; and directing themicroparticles towards the region of nail tissue surface, therebycausing the microparticles to penetrate the nail tissue surface; andscissioning the nail tissue with the impinging microparticles, therebyforming a plurality of free nail microtissue particles, and therebyforming a microconduit through the nail tissue.

[0022] Another embodiment of the invention includes a method fortreating an infection of a tissue underlying nail tissue including thesteps of: accelerating a plurality of microparticles to a velocity thatcauses the microparticles to penetrate into the nail tissue surface uponimpingement of the microparticles on the nail tissue surface; directingthe microparticles towards the region of nail tissue surface; allowingthe microparticles to impinge upon the region of nail tissue surface andto penetrate the nail tissue surface; scissioning the nail tissue withthe impinging microparticles, thereby forming a plurality of free nailmicrotissue particles, and thereby forming a microconduit through thenail tissue; and then administering at least one therapeutic molecule orion by directing the therapeutic molecule or ion into at least onemicroconduit, thereby delivering the therapeutic molecule or ion throughthe nail tissue.

[0023] Another embodiment of the invention includes a method for markingnail tissue with at least one identifying mark or at least onedecorative mark including the steps of: accelerating a plurality ofmicroparticles to a velocity that causes the microparticles to partiallypenetrate into the nail tissue surface upon impingement of themicroparticles on the nail tissue surface; directing the microparticlestowards the region of nail tissue surface; allowing the microparticlesto impinge upon the region of nail tissue surface and to partiallypenetrate the nail tissue surface; scissioning the nail tissue with theimpinging microparticles, thereby forming a plurality of free nailmicrotissue particles, and thereby forming a microconduit through thenail tissue; and then directing a dye or an ink into at least onemicroconduit that partially penetrates the nail tissue, thereby markingthe nail tissue.

[0024] Another embodiment of the invention includes a method forinserting at least one wire through at least one microconduit,including: accelerating a plurality of microparticles to a velocity thatcauses the microparticles to penetrate into a region of nail tissuesurface upon impingement of the microparticles on the nail tissuesurface;

[0025] directing the microparticles towards the region of nail tissuesurface, thereby causing the microparticles to penetrate the nail tissuesurface; scissioning the nail tissue with the impinging microparticles,thereby forming a plurality of free nail microtissue particles, andthereby forming a microconduit through the nail tissue; and directing awire into at least one microconduit, thereby inserting the wire throughthe microconduit. In this embodiment, the microconduit is through thenail tissue where the nail has grown beyond the nail bed and extends outbeyond all other tissue, as in a cantilever or overhang beyond thefinger or toe. In one embodiment, an ornament or jewelry may be attachedto the wire inserted in the microconduit.

[0026] The invention also relates to a method of reducing pressurecaused by a pool of blood beneath an injured or traumatized nailcomprising the steps of: accelerating a plurality of microparticles to avelocity that causes the microparticles to penetrate a region of nailtissue surface upon impingement of the microparticles on the nail tissuesurface; directing the microparticles towards the region of nail tissuesurface, thereby causing the microparticles to penetrate the nail tissuesurface; scissioning the nail tissue with the impinging microparticles,thereby forming a plurality of free nail microtissue particles, andthereby forming a microconduit through the nail tissue; and therebyreleasing the pressure through the microconduit.

BRIEF DESCRIPTION OF THE DRAWINGS

[0027]FIG. 1 is a schematic cross-sectional illustration of anembodiment of the invention of a method and an apparatus for makingmicroconduits in tissue, wherein microparticles are viewed as impingingon a mask containing one or more microholes that expose localizedregions of the skin surface.

[0028]FIG. 2 is a schematic cross-sectional drawing of a collimated beamof microparticles according to one embodiment of the invention impingingonto a region of skin surface.

[0029]FIG. 3 is a schematic cross-sectional representation of anembodiment of the invention including a method and apparatus forelectrophoretic transport of ions through a microconduit and into theepidermis.

[0030]FIG. 4A and 4B are optical photomicrographs of microconduitsformed according to an embodiment of the invention at six sites in theforearm of a subject individual, Subject B.

[0031] FIGS. 5A-5K include confocal images at different depths,indicated in μm from the approximate surface of the skin, of a forearmmicroconduit formed according to an embodiment of the invention inSubject B.

[0032] FIGS. 6A-6I include confocal images at different depths,indicated in μm from the approximate surface of the skin, of a forearmmicroconduit formed according to an embodiment of the invention in asubject individual, Subject A.

DETAILED DESCRIPTION OF THE INVENTION

[0033] The present invention relates to methods and devices foraccelerating microparticles to a velocity that causes a plurality ofmicroparticles to penetrate into a tissue surface upon impingement ofthe microparticles on the tissue surface; directing the microparticlestowards an identified region of tissue surface; allowing themicroparticles to impinge upon the region of tissue surface and topenetrate the tissue surface; scissioning the tissue with the impingingmicroparticles, thereby forming a plurality of free microtissueparticles, and thereby forming a microconduit. As the term is usedherein, “tissue” may include any collection of cells, for example, skincells, including the stratum corneum and the epidermis; nail tissue,including toe nails and finger nails; and muscle cells.

[0034] As used herein, the term “microparticle” refers to a solidparticle that has an approximate diameter or characteristic lineardimension in a range of between about 0.1 (one tenth) micrometer andabout (three hundred) micrometers, or in a range of between about 1(one) micrometer and about 100 (one hundred) micrometers. In oneembodiment, the microparticle has a dimension of about 10 micrometers.In another embodiment, the microparticle has a dimension of about 100micrometers. In yet another embodiment, microparticles having an averagediameter of about 50 micrometers are used to produce a microconduit of adiameter sufficiently large to carry blood and chemicals.

[0035] As used herein, the term “microconduit” refers to a smallopening, channel, or hole into, or through, a tissue, that allowstransfer of materials by liquid flow, and by electrophoresis, themicroconduit being formed upon impact of a plurality of microparticleson the surface of the tissue. In one embodiment, a microconduit may alsoallow materials to move through by diffusion or by convection.

[0036] The average size of a microconduit according to an embodiment isabout one (1) mm or less diameter. In one embodiment, a microconduit hasa diameter in the range of between about 10 micrometers and about 200micrometers. In another embodiment, a microconduit has a diameter in therange of about between 2 micrometers and about one (1) mm. In anembodiment of the invention, a microconduit is usually smaller thanneedles used for syringe injections, but has a characteristic size ordiameter that is much larger than the diameters of carriers of analytemolecules, therapeutic molecules and ions, or the diameters of analytemolecules, therapeutic molecules and ions themselves. As used herein,the term “diameter” refers to the approximate diameter or characteristiclinear dimension of at least one cross-section of an approximatelycylindrical-shaped section of a microconduit. The term “diameter” isalso used to refer to the approximate diameter or characteristic lineardimension of a microparticle or of a molecule.

[0037] The microconduit generating process includes using random shapedmicroparticles of a hard material for impingement on a tissue. In oneembodiment, the major dimension of the particles is about 70 μm. In oneembodiment, particles are accelerated to a velocity in the range of ameter/second, which is far less than a supersonic velocity (300meters/sec). It is presently believed that when the particles strike thetissues of the skin, some of the particles stick to some tissuefragments and bounce off, carrying tissue fragments away; some particlesheat some tissue to a temperature that disrupts the chemical bonds, andthe molecules are reduced to their components of oxygen, hydrogen,nitrogen, and carbon mono or dioxide, which diffuse into the atmosphere,and yet other particles dislodge some tissue fragments by momentumtransfer, ‘knocking’ them away from the locus of the microconduit. Thus,the microconduit formation may be a process that disassembles the skincomponents in tiny increments through various means and removes them bymeans of the gas or liquid flow that accelerates and carries theparticles. The result is an open hole, reasonably circular in crosssection throughout its total length. This has been clearly andrepeatedly shown in the confocal microscope images taken ofmicroconduits. See, for example, the images in FIG. 4A through FIG. 6I.

[0038] Locating a Tissue Surface Region

[0039] According to one embodiment of the invention, the first step increating one or more microconduits into a tissue includes locating asuitable tissue surface region. In the case of a tissue exposed duringsurgery, the location would typically be determined by the skilledinspection of the tissue by the surgeon. For example, if microconduitswere to be formed in cardiac tissue for purposes of introducingangiogenic molecules, the surgeon would locate the appropriate surfaceregion of the heart. In another example, if a dermatological procedure,such as treatment of a skin lesion or condition is desired, theappropriate region of skin surface can be determined through itstopography by a dermatologist. If transdermal drug delivery is desired,after being given general instruction, in many cases the patient candetermine the particular region of skin surface (topographic region) tobe used. In another example, if neonatal intensive care is desired, theattending physician or nurse would locate an appropriate cutaneous orskin surface region on an infant.

[0040] Acceleration of Microparticles

[0041] To form microconduits in tissue using microparticles, themicroparticles are accelerated to a velocity such that their energycauses them to penetrate a region of tissue surface tissue uponimpingement of the microparticles on the tissue surface.

[0042] According to one embodiment of the invention, microparticles areaccelerated to a velocity that causes the microparticles to penetrate atissue surface upon impingement of the microparticles on the tissuesurface, and to scission or microscission the tissue, formingmicroconduits in tissue by using gas flow, in which the gas entrains themicroparticles during the microparticle acceleration, thereby creating aflux of microparticles. As the microparticles approach the tissuesurface, the gas can be preferentially expanded to obtain a significantvelocity component parallel to the tissue surface, thereby minimizinggas entry into the microconduit. If the gas velocity perpendicular tothe tissue surface is too large, gas can enter the tissue. The result,for deep microconduits, of gas entering the tissue, can be undesireddelivery of gas deep into tissue. This problem is avoided by using gasflow that accelerates microparticles while the microparticles aredistant from the tissue surface. If the microparticles are sufficientlymassive, in terms of their volume and mass density, the microparticlemomentum carries the microparticles into the tissue while the gasacquires enough parallel velocity that the gas does not enter theforming microconduits in significant amounts.

[0043] Gas flows suitable for use in an embodiment of the invention canbe obtained in a number of ways. In one embodiment, by usingover-pressure, in which the driving pressure for the gas flow is greaterthan one atmosphere absolute, the flowing gas can be used to acceleratethe microparticles to a sufficient velocity to form microconduits onimpingement on the tissue surface. For example, an over-pressuredreservoir of gas such as a commercially supplied air cylinder can beused.

[0044] Alternatively, in some embodiments of the invention, a partialvacuum can be used to accelerate gas. A partial vacuum is sometimesreferred to as a negative pressure, because it is a pressure less thanthat of one atmosphere absolute. A partial vacuum or suction creates thenegative pressure. In an embodiment utilizing a partial vacuum to creategas flow, the risk of incidental delivery or injection of the flowinggas into the tissue in which one or more microconduits are being formedis reduced. In this case, the skin surface is exposed to less than oneatmosphere. As a result, the possible injection of gas into the skin isless likely than it is when the gas flow is driven by a positive(greater than one atmosphere). To create a partial vacuum for use in anembodiment of the invention, the venturi effect can be used. Accordingto an embodiment, to accelerate microparticles dispersed within aseparate container, the container is kept at a pressure greater than theventuri pressure (usually atmospheric pressure). The gas flowing past anopening or orifice of the container then has a reduced relativepressure, and microparticles are thereby caused to move out of theseparate container into the gas flow. This method is well known to thoseskilled in the art as the basis for relatively inexpensive microscissiondevices, such as an “air brush.”

[0045] In one embodiment, because of its ready availability and lowcost, either pressurized or ambient air, at about one atmosphereabsolute, may be used as the microparticle-accelerating, flowing gas.The air should be either dry, dehydrated, or of low-moisture content.This is necessary because microparticles can clump together if theirsurfaces have water molecules on them. In one embodiment, nitrogen gasfrom a cylinder of compressed nitrogen or from a container of boilingliquid nitrogen is used. In one embodiment, an over-pressured reservoirsuch as a commercially supplied air cylinder can be used. In otherembodiments, examples of gases suitable for use to acceleratemicroparticles include inert gases such as argon, either at normalatmospheric pressure, or at a higher or lower pressure than oneatmospheric absolute.

[0046] A refrigerant gas, for example, 1,1,1,2 tetrafluoroethane, thatsignificantly cools as it is expanded can also be used to acceleratemicroparticles. The temperature of 1,1,1,2 tetrafluoroethane drops to−60 degrees F. when released from a spray can. This is important ifmicroparticles, such as ice or solid water, that subsequently meltwithin the tissue are used. In another embodiment, cooling of the tissuesurface is used to alter mechanical and physical properties of thetissue before and/or during microparticle impingement. In oneembodiment, cooling of the tissue surface occurs because the flowing gasused to accelerate the microparticles is at a temperature of below about20° C.

[0047] In yet another embodiment, the microparticles are accelerated bymeans of a flowing liquid. In a particular embodiment, the flowingliquid is at a pressure greater than about one pound per square inch. Inone embodiment, the temperature of the flowing liquid is below about 20°C.

[0048] According to another embodiment of the invention, microparticlesare accelerated by impact with a moving solid surface that is moving ata sufficiently high velocity. One method for achieving a suitable movingsolid surface is to use a rapidly rotating impeller. For example, inorder to accelerate the microparticles, they can be drawn out of acontainer by means of evaporation or by reduced pressure, or pushed outby a positive pressure, and contacted with a rapidly rotating, hardsurface.

[0049] Tissue Microscissioned or Compacted

[0050] According to an embodiment of the invention, microparticles areconstrained to impinge mainly onto localized regions on the tissuesurface, with such force that tissue material is removed by momentumtransfer, “scissioned” or “microscissioned” from the tissue upon impactof the microparticles with the tissue. As used herein, the terms“scissioned,” “microscissioned,” “microscission,” and grammaticalvariations thereof, refer to an opening up of the tissue, and theprocess of scission, microscission or opening up of tissue may involveremoving part of the tissue by rendering tissue into gases or liquids,or by attachment of microparticles to parts of tissue and carrying themoff by momentum transfer.

[0051] The process of scission or microscission of tissue, for exampleskin or nail tissue, according to an embodiment of the invention is aprocess of cutting, or microcutting, or cleaving the tissue, therebyforming a plurality of free microparticles of tissue, and therebyforming a microconduit. As the microparticles and air are forced intothe tissue, the air turns around and carries out a majority of the newlyformed microtissue particles and the impinging microparticles, therebyforming a microconduit in the tissue. The tissue is disassembled in amechanical way by impinging microparticles.

[0052] In one embodiment, the tissue that is microscissioned is ejectedfrom the tissue surface. Although a theoretical understanding of theinteraction of impinging microparticles with tissue is not required topractice the invention, it is believed that the incident angle of themicroparticle and the velocity may be important. During the initialstage of microconduit formation, when the forming microconduit depth issmall, tissue material may be ejected from the forming microconduit. Inthe later stages of microconduit formation, when microconduit depth isgreater, some cells of the tissue may be damaged, and the tissue may beprogressively compacted, such that smaller amounts of tissue materialare ejected from the microconduit than in the initial stages ofmicroconduit formation. In another embodiment, microscission of tissuemay relate to an opening up of tissue by compacting the tissue uponimpingement of the microparticles on the tissue.

[0053] It is true that the deeper a microconduit is made, the moredifficult it is for the incoming gas/particles to flow to the bottom ofthe dead end cylinder of the microconduit and for the spent particlesand tissue particles to flow back upstream and exit the microconduit.Thus, the microconduit generation rate will diminish with depth and maypossibly be used to predetermine the microconduit depth reached bycontrolling the diameter of the mask microhole and thus the diameter ofthe microconduit.

[0054] Microparticles Used

[0055] In one embodiment of the invention, solid, persistentmicroparticles that do not dissolve within the tissue after impinging,such as microparticles comprised of aluminum oxide, also referred to as“alumina,” are used to form suitable microconduits.

[0056] Another class of microparticles suitable for use in an embodimentinclude solid phase microparticles comprised of biocompatible substancesthat exist in the liquid state at normal physiologic tissuetemperatures, for example normal human body temperature, which is about37 degrees Celsius, in the interior, and often lower at the skin'ssurface. In one embodiment, the microparticles have a melting point lessthan about 33 degrees Celsius. Such solid phase microparticles impingeonto a localized region of tissue surface, and enter into the tissue,creating microconduits. Such solid phase microparticles melt within thetissue, and the resulting liquid mixes with tissue interstitial fluid.The change to the liquid state further results in diffusion and removalof the microparticle material. Accordingly, in one embodiment,microparticles comprised of solid water (water ice) or other tissuesoluble material are used. In another embodiment, use of microparticlesthat are water soluble and dissolve within a tissue after impingementalso has the advantage that the particles do not persist within thetissue. It is important, however, to use microparticle materials that donot cause significant localized irritation or localized tissue damagewhile dissolving. This can be accomplished by choosing microparticlematerials that have an acceptable combination of lack of irritation andrapid diffusion of soluble molecules away from the site of themicroparticle in the tissue.

[0057] In one embodiment, microparticles comprised of soluble,biocompatible substances may be used to form microconduits. In aparticular embodiment, microparticles may include sodium bicarbonate,which readily dissolves in water within a tissue, particularly theepidermis. Sodium bicarbonate occurs naturally in the body, and isessentially nontoxic. Temporarily elevated concentrations of bicarbonatecan cause temporary local pH increases, but these are not believed tocause significant side effects. If such a pH increase is undesirable, inone embodiment microparticles comprised of a buffering mixture of sodiumacetate and sodium bicarbonate can be used. Upon dissolution of suchbuffered microparticles, only an insignificant pH change from the normalepidermal pH will occur. In yet other embodiments, microparticles arecomprised of other nontoxic, suitable salts such as lactates, sterates,and the like. Urea is another naturally occurring substance that isrelatively nontoxic. Accordingly, in another embodiment, microparticlesare comprised of urea or urea salts.

[0058] In another embodiment, microparticles include solid phasemicroparticles comprised of biocompatible substances that exist in thegaseous state at normal physiologic tissue temperatures. In a particularembodiment microparticles are comprised of solid phase carbon dioxide,also known as “dry ice.” As is well known through use of carbon dioxidein fire extinguishers, solid carbon dioxide can be formed by rapidexpansion from the liquid phase. After impingement and entry intotissue, solid carbon dioxide microparticles of appropriate size sublimerapidly within the tissue and thereby cause a pressure burst. Theimpinging solid carbon dioxide microparticles first contribute tomicroconduit formation, then undergo an almost explosive burst that alsocontributes to formation of the microconduit, as gaseous carbon dioxiderapidly forms. The pressure of the expanding gas assists microconduitformation because the rapid sublimation of impinging carbon dioxidemicroparticles causes an almost explosive release of gas within theforming microconduit. However, if solid carbon dioxide is used, caremust be taken if a significant number of carbon dioxide microparticlesremain in the tissue, because a several order of magnitude increase involume occurs as the solid carbon dioxide sublimes to form gaseouscarbon dioxide.

[0059] In one embodiment, solid phase microparticles contain solutes,including one or more therapeutically effective substances or drugs, andserve to deliver drugs into the skin upon subsequently melting. As usedherein, the terms “therapeutically effective substance” or “therapeuticsubstance” or “drug” include:

[0060] (i) Compounds and compositions recognized in the official UnitedStates Pharmacopoeia, the official Homeopathic Pharmacopoeia of theUnited States, or the official National Formulary, or any supplement ofany of them;

[0061] (ii) Compounds and compositions intended for use in thediagnosis, cure, mitigation, treatment, or prevention of disease in manor other animals; and

[0062] (iii) Compounds and compositions (other than food) intended toaffect the structure or any function of the body of man or otheranimals.

[0063] In another embodiment, inert microparticles, such as ceramiccarriers or polymeric carriers, may be used to both form microconduitsand to serve as slow-release devices.

[0064] An Apparatus for Impinging Microparticles

[0065] This invention also relates to an apparatus for use in creatingone or more microconduits by microparticle impingement. In oneembodiment, the apparatus includes a means for accelerating a pluralityof microparticles to a velocity that causes the microparticles topenetrate into a surface of the tissue upon impingement of themicroparticles on the tissue surface; a means for directing themicroparticles towards a region of tissue surface; and a means forallowing the microparticles to impinge upon the region of tissue surfaceand to penetrate the tissue surface, thereby forming a microconduit inthe tissue. Thus, the apparatus can create a flux of microparticles thatis concentrated, and that impinges on one or more small, localizedregions of a tissue.

[0066] Controlling the Amount of Microparticles and Controlling theRegion of Tissue Surface Impinged by Microparticles

[0067] In one embodiment, the amount of microparticles that are allowedto impinge upon a region of tissue surface is controlled by controllingthe area of the region of tissue surface exposed to incoming,accelerated microparticles.

[0068] According to an embodiment of the invention, localized areas oftissue surface can be defined by use of a mask comprising a membraneplaced on, at, or near the tissue surface, the mask containing one ormore microholes. In this embodiment, microparticles directed towards themask impinge onto a localized area of a tissue surface region if theypass through a microhole in the mask. In one embodiment, the microholeshave a diameter in a range of about three micrometers and about onethousand micrometers. According to an embodiment, a suitable mask can befabricated from a membrane (a thin sheet of material that typically hasan area of several square millimeters or more). The mask preventsmicroparticles from passing through the mask material. As used herein,the term “membrane” refers to a small sheet of material that is muchlarger in lateral extent than the localized areas onto whichmicroparticle impingement is sought. According to an embodiment, themembrane has at least one microhole that defines the local tissuesurface area when the mask is held against, or next to, a tissuesurface. Much of the tissue surface is thereby protected frommicroparticle impingement, such that localized surface areas are definedby the restriction that microparticles can only reach the tissue if themicroparticles are incident within a microhole. In an embodiment inwhich a mask is used in forming a microconduit, generally themicroconduit opening at the tissue surface is smaller than themicrohole.

[0069] In one embodiment, the membrane may be conformable to the tissuesurface.

[0070] According to an embodiment, suitable masks may be formed fromsheets made of solid materials such as metals, metal alloys, polymers,silicon, passivated silicon, ceramics and glasses. In a particularembodiment, a mask may be comprised of a solid biocompatible materialsuch as titanium, stainless steel, polyimide, nylon, poly(lactic-co-glycolic acid), calcium aluminum phosphate, and the like.Depending on the method used to accelerate the microparticles, masks ofa variety of thicknesses may be suitable for use in an embodiment. Inone embodiment, a mask has a thickness in the range of about one (1)micrometer to about one thousand (1,000) micrometers (μm). In the caseof gas-accelerated microparticles, it is presently preferred to usemasks with a thickness of about twenty five (25) to about one hundred(100) μm. In one embodiment, a mask has a thickness of about 50micrometers. In another embodiment, a mask has a thickness of about 75to about 100 micrometers. A mask as thin as perhaps one micrometer couldbe used if it didn't wear away by the impinging particles before thedesired microconduit was generated. Also, a 1 micrometer membrane, evenif it continued to mask for the 4 to 14 seconds required to form amicroconduit, would likely not have the stiffiess to stay tightlyagainst the stratum corneum. Masks with multiple microholes tend to‘billow up’ away from the stratum corneum, and must be stiff enough tocompletely resist this gas-driven ‘sail’ phenomenon. As is well known tothose skilled in the art of semiconductor microfabrication, a variety ofestablished techniques can be used to form microholes in a mask.Suitable techniques include use of rotating drills, use of laser energy,and spatially constrained chemical, plasma, or reactive ion, or ionetching.

[0071] According to an embodiment, the fabricated mask can be heldagainst a tissue by a variety of methods. In the case of transdermalmicroconduit formation, one embodiment comprises holding a mask mountedon a holding fixture which also holds and aligns the gas/particleemitting nozzle against the skin, such as the forearm, by using anelastic strap, similar to a snugly fitting wrist watch. In a particularembodiment of the invention, formation of microconduits through thestratum corneum typically requires 4 to 20 seconds of microparticleimpingement. A short time is used to put on the mask/strap, so that thetotal time of holding the mask firmly against the skin is at most a fewminutes.

[0072]FIG. 1 is a schematic, cross-sectional illustration of oneembodiment of the invention of a method and an apparatus for makingmicroconduits in tissue. In this embodiment, the tissue comprises thestratum corneum 18. In FIG. 1, microparticles are viewed as impinging ona mask 10 disposed on the stratum corneum 18 of skin. Mask 10 definesone or more microholes that expose localized regions of the skinsurface. The plurality of irregular-shaped objects (12, 12″, 12, 12′″)from which arrows 14, 16 eminate represent impinging microparticles. Themask or micromask material 10 blocks microparticles 12′ through 12′″from reaching the surface of the tissue. The microparticles 12′ through12′″ bounce off the mask material in a direction shown generally byarrow 14, without reaching the stratum corneum 18, or the epidermis 20.The microparticles 12 traveling in a direction shown generally by arrow16 pass through the microholes in the mask and impinge upon the surfaceof the stratum corneum 18. It should be noted that in a preferredembodiment the angle at which microparticles impinge upon a tissue,including the stratum corneum, is close to ninety (90) degrees. Theangle of impingement may be controlled, in different embodiments, bycollimating the microparticles, as described below.

[0073] In order to define one or more localized areas for microparticleimpingement, one or more microholes with microhole size (diameter ifessentially circular) in a range of between about three (3) micrometers(μm) and to about one thousand (1,000) micrometers (μm) are provided inthe membrane. The mask is held snugly against the tissue surface region.Incident microparticles are thereby constrained to impinge only onto thelocalized tissue areas exposed through the microholes in the mask.According to an embodiment of the invention, it is necessary that themicroholes be somewhat larger than the microparticles, and therefore,the opening of the microconduit formed in tissue is generally smallerthan the microholes in the mask.

[0074] Any of several attachment or positioning means are satisfactoryfor temporarily holding the mask against a tissue surface region. In oneembodiment, an adhesive backing can be provided on the side of the maskthat is to contact the tissue surface region. Pressure contact can alsobe used, for example, by affixing a strap that goes around the body orbody part such that the mask is held against the tissue. For example, ifthe tissue surface region is the skin of the forearm, a strap similar tothat used on a wrist watch can be used. In one embodiment, a mask mayalso be incorporated into a device that contains the source ofmicroparticles, such that placing the device against a tissue allowsmicroparticles to be impinged onto localized areas in order to make oneor more microconduits.

[0075] According to another embodiment, a region of tissue surface isdefined by directing a collimated beam of microparticles onto the regionof tissue. In a particular embodiment, the size (for example, thediameter, if a cross-section of a circular beam is used) of themicroparticle beam is the main determinant of the diameter of theresulting microconduits.

[0076] The idea of collimating the particle stream is that, in aperfectly collimated column, all particles would be moving parallel toone another and perpendicular to the surface upon which they impinged,assuming that the stream itself was perpendicular to the surface. Thus,the diameter of the surface upon which the particles impinged would bethe same as the diameter of the particle stream. Normally, thecomponents of a particle-loaded gas stream would diverge, growing largerin diameter the greater the distance from the particle source. Aperfectly collimated particle stream would not diverge, and if thestream diameter could be 150 micrometers, no mask would be needed, themicroconduit would be 150 micrometers in diameter also. Perfectcollimation is impossible to achieve, so a mask, using a well collimatedstream, is required to more precisely define the microconduit diameterat the surface of the stratum corneum. However, if the mask is not tightagainst the stratum corneum, the stream will begin diverging as itleaves the underside of the mask. With rather good collimation, thiseffect is small, and every microhole in the mask does not need to betight against the stratum corneum, because even though it isn't the beamsize, it is close to the mask microhole diameter.

[0077] One way to collimate the stream is to use a nozzle with a longbore of the desired diameter. FIG. 2 shows another way, where only theends of a long bore exist. In FIG. 2, a plurality of masks, generallyindicated by 40, includes mask 10 and mask 24. An incoming, slightlydivergent particle stream hits the top of the mask 10, which filters outthe most divergent particles. The better collimated particles then gothrough the microholes in that mask, and diverge only slightly beforereaching the lower mask 24, which filters out those particles thatdiverge too much. The particles exiting mask 24 are well enoughcollimated that mask 24 does not need to be held tightly against thestratum corneum, 18 in order to produce microconduits nearly the samesize as the microholes in mask 24.

[0078] In FIG. 2, at least two sheets of material (10, 24) arepositioned so as to substantially prevent microparticles (12′″) fromreaching some areas of the tissue, with the result that microparticles12′″ strike the mask and bounce off in the direction shown by arrow 22,for example. Impinging microparticles (12), traveling in the directionshown by arrow 16, predominantly strike one or more localized areas ofthe tissue surface. FIG. 2 shows that the use of at least two sheets ofmask material can serve to approximately collimate the impingingmicroparticles according to an embodiment of the invention.

[0079] In one embodiment, the amount of microparticles that are allowedto impinge upon a region of tissue surface is controlled by controllingthe amount of time that microparticles are allowed to impinge on theregion of tissue. This is true whether or not a collimated microparticlebeam is used. In one embodiment, the time of microparticle impingementonto tissue surface, and the amount of microparticles impinging on aregion of tissue surface can be controlled by scanning the beam over theregion of tissue surface, and by using both the scan rate (velocityacross or parallel to the tissue surface) and the time the beam is on oroff. According to an embodiment, the on/off time can be controlled bygating the beam, for example, by moving a deflecting solid surface outof or into the beam.

[0080] In one embodiment, control of the microparticle amount can beaccomplished by an indirect method in which the microparticle beam iscalibrated in separate measurements, such that timing of the “ON” timethereafter controls the amount of microparticles impinging onto alocalized surface area. Here, a test material in which a microconduitcan be easily measured is subjected to the particle beam for a fixedamount of time. The resulting ‘test’ microconduit depth is measuredmicroscopically. By having performed calibration experiments relatingthe microconduit depth in the test material to the microconduit depth ina human subject's skin, an exposure time can be elucidated to generatethe desired microconduit depth in the human subject.

[0081] The number of microparticles per unit volume in a gas stream of agiven diameter and velocity can be varied. This can be done by meteringthe particles from the particular reservoir into the gas stream through:a) changing the aperture diameter and/or the number of apertures throughwhich the particles fall from the particle reservoir into the passinggas stream, or b) changing the rate and/or amplitude at which theparticle reservoir is shaken to dislodge the particles and bring themacross the aperture to fall into the gas flow.

[0082] In a particular embodiment, a particle flux which is generallyused to produce a microconduit is about one million particles persecond. Based on a calibrated count and weight of particles, the numberof particles per second transmitted in a flux can be determined throughthe time/weight relationship.

[0083] The same is true with the delivery of particles by the ‘impeller’approach. The number of particles per unit time being hit by theimpeller can be varied by the number of particles available to be hit.

[0084] Generally, by impinging microparticles onto a localized areaequal to a range of about between about 100 square micrometers and aboutone million square micrometers (for example, an area with a diameter ofabout five to about one thousand micrometers (μm)), microconduits withapproximately the same cross-sectional size can be formed. In oneembodiment, the microparticles impinge upon the region of tissue surfacewhich has an area in a range of between about 100 square micrometers andabout two million square micrometers.

[0085] In the case of skin, according to one embodiment, microparticlesare impinged onto a localized area equal to between about 1000 squaremicrometers and about 100,000 square micrometers (for example, an areawith a diameter of about 30 to about three hundred micrometers), to formmicroconduits of approximately the same cross-sectional size.

[0086] The present invention also relates to an apparatus suitable forformation of microconduits in tissue by microparticle impingement. Inone embodiment, an apparatus includes a scannable microparticle beamcollimator with an on/off gating means; a source of movingmicroparticles; at least one solid surface with at least onebeam-defining aperture; a means for gating the microparticle beambetween “ON” and “OFF;” a first excess particle collection duct andsuction means, for recovering microparticles that do not pass throughthe beam-defining aperture; and a second excess particle collection ductand suction means, for recovering microparticles that pass through thebeam-defining aperture, impinge onto the tissue, and rebound from thetissue. In one embodiment the beam-defining aperture of the apparatusis, for example, in the size (diameter) range of about between five (5)micrometers (μm) and about two thousand (2,000) micrometers (μm); inanother embodinent the aperture size is in the range of between aboutthirty (30) micrometers and about one thousand (1,000) micrometers. Inanother embodiment, the apparatus can comprise more than two solidsheets provided with at least one collimating aperture, to furthercollimate the microparticle beam.

[0087] According to an embodiment, the depth of the microconduits formedby microparticle impingement can be regulated by controlling the amount(number or total mass) of the impinging microparticles, the size(individual mass) of impinging microparticles, the speed (magnitude ofthe microparticle velocity), the angle of impingement (direction of themicroparticle velocity) the hardness of the microparticles, the shape ofthe microparticles, the sharpness (cutting ability) of themicroparticles, the total number or amount of impinging microparticlesthat impact a local surface area, the mask geometry (microhole size andmask thickness), and in some cases, the mask material (which can governthe degree of elasticity and the amount of scattering of microparticlesfrom the walls of the microholes of the mask). The viscosity andvelocity of any gas between the microparticle source and the local areaonto which microparticles impinge can also affect microconduitformation.

[0088] Humidity Depth Sensors

[0089] According to an embodiment, the formation of microconduits andthe depth of the microconduits formed by microparticle impingement canbe guided in whole, or in part, by sensing or measuring the humidity ofthe gas that is returned from the forming microconduit. For example, thestratum corneum and the nail plate of finger and toe nails have a lowwater content compared to viable underlying tissue. A dry gas can beused to accelerate and impinge the microparticles on a microlocalizedarea of the skin or nail. Gas that enters and then returns from amicroconduit can acquire water according to the wetness of the tissueinto which the microconduit penetrates. A microconduit that does notpenetrate the stratum corneum (or nail) will transfer less water to thegas.

[0090] The gas flow can be essentially continuous, or if it is desired,the gas flow can be stopped or interrupted (modulated) to increase itswater content. Some of the gas which flows away from the microconduitsite can be monitored for humidity, using humidity sensors, such aselectrical capacitance-based humidity sensors. Examples of the generalprinciple of such humidity sensing can be found in Schiffinan et al.,“Airway humidification in mechanically ventilated neonates and infants:a comparative study of a heat and moisture exchanger vs. a heatedhumidifier using a new fast-response capacitive humidity sensor,” Crit.Care Med. 25:1755-1760 (1997); and Ohhashi et al., “Human perspirationmeasurement,” Physiol. Meas. 19: 449-461 (1998), the teachings of whichare incorporated herein by reference in their entireties.

[0091] Modification of Tissue Surfaces

[0092] According to an embodiment, the general process of the inventioncan also be carried out by including the additional step of applying oneor more conditions at the surface of the tissue in order to alter themechanical properties of the outer layer of the tissue surface. Forexample, in one embodiment, the interaction of impinging microparticleswith the outer layer can in some cases be varied by changing thehardness of the outer layer. In particular, it may be noted thatimpinging microparticles tend to scission a harder surface, and thenbounce off after impingement, also removing parts of the harder surface.In contrast, if microparticles impinge onto a softer surface,microparticles tend to penetrate into, and then reside, within thetissue. Accordingly, because many tissues become more rigid as thetemperature is decreased, in one embodiment, cooling may be used toincrease the hardness of a tissue surface layer. Thus, applying acooling stream of gas or liquid to the tissue before microparticleimpingement can be used in an embodiment to harden the tissue surface.In the case of skin, cooling of the skin's surface before impingement bymicroparticles may decrease sensation.

[0093] In the case of the skin, the outer region of the nonviablestratum corneum layer is often significantly harder than the viableepidermis. If, however, the skin's surface is placed in contact withwater, or is occluded such that exiting water accumulates within thestratum corneum, the stratum corneum becomes softer. Accordingly, oneembodiment includes the step of applying a desiccating material thathardens the stratum corneum by removing water. Suitable materials foruse in an embodiment include isopropyl alcohol, ethanol, and otheralcohols. In a particular embodiment, drying the stratum corneumpartially by a dry gas stream, such as dry air, can also be used.

[0094] According to an embodiment, after a microconduit is formed,recovery processes begin that eventually lead to full restoration of theskin's barrier function. As the term is used herein, “recovery” of amicroconduit refers to the self-repair process in which the tissue thathas been microscissioned away is replaced by newly grown tissue. Onecommon feature of recovery is the re-establishment of lipid membranes,which allow passage of nonpolar molecules, but exclude ions and chargedmolecules. In the human skin approximately one hundred (100) lipidbilayers must be penetrated to pass completely through the stratumcorneum. In one embodiment, following formation of a microconduit, thepermeability of the skin at the site of the microconduit can beincreased by applying one or more electrical pulses. The electricalpulses applied cause electroporation of at least one lipid-containingmembrane. In a particular embodiment, the newly formed microconduit isallowed to recover for a period of time before the electrical pulse isapplied. As used herein, the term “partially recovered” as it applies toa microconduit, means that the tissue, removed or damaged by impingingmicroparticles that formed the microconduit, has begun to repair itselfor to grow new tissue. Although fifty (50) to three hundred (300) voltstypically are needed to achieve electroporation of human skin that hasnot been subject to a process for formation of one or moremicroconduits, the voltage of an electrical pulse can be less than 50volts to achieve microlocalized electroporation of skin that has alreadyundergone microconduit formation, and undergone partial recovery.

[0095] A partially recovered microconduit is a site that preferentiallyexperiences electroporation, because a voltage across the stratumcorneum associated with either an electrical current pulse (“currentclamp” pulse) or an electrical voltage pulse (“voltage clamp” pulse)will concentrate mostly across fewer lipid membranes at the site of apartially recovered microconduit. For example, if approximately five (5)lipid bilayer membranes have been formed within or near the epidermalentrance to a stratum corneum-penetrating microconduit, then a pulseneeds to cause only approximately five (5) V across the partiallyrecovered microconduit. This is much less than the fifty (50) to threehundred (300) V associated with localized transport regions (LTRs)caused by electroporation in human skin not subjected to microconduitformation processes. For a description of such localized transportregions see, for example, Pliquett, et al “Imaging of FluorescentMolecules and Small Ion Transport Through Human Stratum Corneum DuringHigh-voltage Pulsing: Localized Transport Regions are Involved,” 58 J.Biophys. Chem.,185-204, 1996; Prausnitz et al. “Imaging Regions ofTransport Across Human Stratum Corneum During High Voltage and LowVoltage Exposures,” 85 J. Pharm. Sci. 1363-1370, 1996; Weaver, et al.“Theory of electrical formation of aqueous pathways across skintransport barriers,” 35 Advanced Drug Delivery Reviews 21-39, 1999; andGowrishankar et al., “Spatially Constrained Localized Transport RegionsDue to Skin Electroporation,” 60 J. Controlled Release 101-110, 1999,the teachings of which are incorporated herein by reference in theirentireties. Although the processes governing formation of LTRs in humanskin subject only to electrical pulses are not fully understood,preferential electroporation at skin sites with lipid membranes isqualitatively consistent with LTR formation.

[0096] In another embodiment, microscopic aqueous pathways associatedwith electroporation of lipid membranes at the site of a partiallyrecovered microconduit are altered by the further step of applying atleast one modifying agent. The modifying agent serves to alter aqueouspathways formed by electroporation, and can be used to additionallyassist the re-opening of a microconduit that has been partially closedoff by natural recovery processes. This embodiment includes theapplication of long, linear molecules that enter aqueous pathways formedby electroporation of lipid-containing membranes, such as cellmembranes, or one or more multilamellar lipid bilayer membranes of arecovering stratum corneum. Suitable modifying agents for use in anembodiment include dextran, heparin, and DNA. Such modifying agents canprolong the lifetime of aqueous pathways through lipid membranes, andcan also alter the size of the aqueous pathways through lipid membranes.This increases the permeability of a partially recovered microconduitand also decreases the electrical resistance of a partially recoveredmicroconduit.

[0097] The invention also relates to a method for altering, and therebycontrolling, the skilys recovery response to formation of one or moremicroconduits. As used herein, the term “recovery” refers to there-growth of tissue removed or otherwise damaged by the formation ofmicroconduits, as to close off and seal the opening of a microconduit atthe tissue surface. An embodiment of the method includes supplying (orremoving) chemical agents at the site of the entrance to one or moremicroconduits, or at a site within microconduits, such that theconcentration of the chemical agents is controlled. Controlling theconcentration of chemical agents within the tissue surrounding amicroconduit can alter the rate of recovery of the tissue and of themicroconduit. Although eventual recovery is generally sought, it can beadvantageous to either cause delay in the recovery (keeping themicroconduit open), or to accelerate recovery, once desired molecularand ionic transport or measurements have been accomplished.

[0098] This method is particularly useful with transdermalmicroconduits. In one embodiment, calcium ion (Ca²⁺), for example, isused as the chemical agent. If Ca²⁺ is present at relatively highconcentrations within the microconduit, then the Ca²⁺ concentrationwithin the epidermis is also high, and repair and recovery processestend to be inhibited, and the microconduits tend to remain open andavailable for molecular and ionic transport. In another embodiment,5-fluorouracil is used to delay or prevent recovery of a microconduit.

[0099] In yet another embodiment, a retinoid such as retinoic acid; asurfactant; or an antigent is used to delay recovery of a microconduit.In a particular embodiment, the chemical agent used to delay repair andrecovery of the microconduit is topically applied to the opening of themicroconduit, directed into the opening, and applied to tissuesurrounding the opening. In a particular embodiment, the chemical agentdirected into the opening is in a column, for example, a pipet orcapillary tube, and the column is sealed to the tissue around themicroconduit. Next, pressure is directly applied to the microconduit,for example, by squeezing a rubber bulb attached to one end of thecolumn or pipet, thereby forcing the chemical agent included in thecolumn or pipet into the microconduit. The chemical agent is absorbed bythe tissue surrounding the microconduit.

[0100] The method for altering, and thereby controlling, the skin'srecovery response to formation of one or more microconduits bycontrolling the concentration of chemical agents within the tissuesurrounding a microconduit is applicable also to microconduits formed byelectroporation and keratolytic agents (see, for example, U.S. Pat. No.5,911,223 to Weaver et al., “Introduction of Modifying Agents into Skinby Electroporation,” June 15, 1999; Zewert et al., “Creation ofTransdermal Pathways for Macromolecule Transport by Skin Electroporationand a Low Toxicity, Pathway-Enlarging Molecule,” Bioelectrochem.Bioenerget. 49:11-20, 1999; Ilic et al., “Spatially Constrained SkinElectroporation with Sodium Thiosulfate and Urea Creates TransdermalMicroconduits,” 61 J. Control. Release,185-202, 1999), the teachings ofwhich are incorporated herein by reference in their entireties.

[0101] The method for controlling the skin's recovery response toformation of one or more microconduits by controlling the concentrationof chemical agents within the tissue surrounding a microconduit isapplicable also to micropores in skin formed by localized laser ablation(see, for example, S. L. Jacques et al., “Controlled Removal of HumanStratum Comeum by a Pulsed Laser,” 88 J. Invest. Dermatol, 88-93, 1987;by ultrasound (see, for example, N. Yamashita et al., “Scanning ElectronMicroscopic Evaluation of the Skin Surface After Ultrasound Exposure,”247 The Anatomical Record 455-461, 1997; T. Hikima et al., “Effect ofUltrasound Application on Skin Metabolism of Prednisolone 21-Acetate,”15 Pharm. Res.,1680 -1683, 1998; J. Wu et al. “Defects Generated inHuman Stratum Corneum Specimens by Ultrasound,” Ultrasound in Med. &Biol. 24:705-710, 1998)), the teachings of which are incorporated hereinby reference in their entireties. This method of altering the recoveryof skin is also applicable to microconduits or skin openings formed by“thermal poration” (see, for example, U.S. Pat. No. 6,142,939 to J. A.Eppstein et al., “Microporation of Human Skin for Drug Delivery andMonitoring Applications;” A. Smith et al., “Fluorescein Kinetics inInterstitial Fluid Harvesting from Diabetic Skin during FluoresceinAngiography: Implications for Glucose Monitoring,” 1 Diabetes Tech.Therapeut.21-27, 1999); by insertion and removal of microneedles (see,for example, S. Henry et al. “Microfabricated Microneedles: A NovelApproach to Transdermal Drug Delivery,” 87 J. Pharm. Sci. 922-925,1998); or by any other means for creating a stratum corneum-penetratingopening less than about five hundred (500) micrometer (μm) in diameteror characteristic length within the opening. The teachings of the abovecited publications are incorporated herein by reference in theirentireties.

[0102] Examples of methods and apparatus used in electroporation; incontrolling transport of molecules across tissue using electroporation;in treatment of cells in a tissue; and in making biopotentialmeasurements, include those which are disclosed in related U.S. Pat. No.5,019,034 to Weaver et al., filed on Mar. 30, 1989, which issued as apatent on May 28, 1991; B1 5,019,034 to Weaver et al., filed on Nov. 9,1993, which issued as a patent on Aug. 15, 1995; U.S. Pat. No. 5,389,069to James C. Weaver, filed Sep. 17, 1993, which issued as a patent onFeb. 14, 1995; U.S. Pat. No. 5,547,467 to Prausnitz et al., filed onJul. 23, 1993, which issued as a patent on Aug. 20, 1996; U.S. Pat. No.5,667,491 to Pliquett et al., filed Jun. 7, 1995, which issued as apatent on Sep. 16, 1997; U.S. Pat. No. 5,749,847 to Zewert et al., filedJun. 6, 1995, which issued as a patent on May 12, 1998; U.S. Pat. No.5,983,131 to Weaver et al., filed Aug. 9, 1996, which issued as a patenton Nov. 9, 1999; U.S. Pat. No. 5,911,223 to Weaver et al., filed Aug. 9,1996, which issued as a patent on Jun. 15, 1999; U.S. Pat. No. 6,085,115to Weaver et al, filed May 22, 1998, which issued as a patent on Jul. 4,2000; and in related U.S. application Ser. No. 60/189,670, filed on Mar.15, 2000; Ser. No. 60/209,985, filed on Jun. 8, 2000; and Ser. No.60/228,488, filed on Aug. 28, 2000. The entire teachings of theabove-referenced patents and applications are incorporated herein byreference in their entireties.

[0103] Other Embodiments

[0104] Other embodiments of the invention relate to microconduits thatpenetrate partially or fully through the skin's stratum corneum, themicroconduits formed by microparticle impingement onto one or morelocalized surface areas of the stratum corneum. According to anembodiment, microconduits that fully penetrate the stratum corneum areof particular interest, because such microconduits provide large aqueouspathways for molecular and ionic transport through the stratum corneum,the skin's main barrier to ionic and molecular transport. Becausemicroconduit size, according to an embodiment, is much larger than evenmacromolecules such as proteins and nucleic acids, transport occurs withinsignificant steric hindrance. For this reason, trans-stratum corneummicroconduits or trans-comeal microconduits can provide transdermaltransport of essentially any size molecule.

[0105] According to an embodiment, suitable transdermal microconduitsthat fully penetrate the skin's stratum corneum can be formed by usingmicroparticles which do not dissolve within the tissue after impinging.In such an embodiment, it is preferred to form microconduits which donot exceed a range of between approximately forty (40)micrometers andsixty (60) micrometers (μm) in depth, as measured from the outer surfaceof the stratum corneum. At this depth the microparticles remain withinthe viable epidermis, and above the stratum basal epidermidis (a layerof stem cells that continually replenishes the epidermis and stratumcorneum). As a result, in approximately two weeks the entire epidermisis replaced and renewed, with epidermal cells differentiating and movingoutward to replenish the nonviable stratum corneum, and carrying thewater insoluble microparticles out of the body. According to anembodiment, although water insoluble microparticles within this depth inthe epidermis do not dissolve in the epidermal tissue, themicroparticles usually do not remain in the tissue for more thanapproximately two weeks. Microparticle materials which do not causeirritation are therefore suitable for creating transdermal microconduitswhich do not exceed approximately forty (40) to sixty (60) micrometers(μm) in depth.

[0106] A present preferred embodiment is to form microconduits less thanabout fifty micrometers (50 μm) in depth by using aluminum oxide(alumina) microparticles that are irregular in size, and that have acharacteristic linear dimension of about thirty to seventy micrometers(30 to 70 μm).

[0107] According to an embodiment, microconduits deeper than fiftymicrometers (50 μm) can also be formed if the microparticle material issufficiently inert. Presently it is preferred to use microparticlescomprised of aluminum oxide (alumina) to form microconduits deeper thanapproximately fifty micrometers (50 μm). In some cases, however, agranuloma can be formed by cells which phacytose such microparticles.Such granulomas may occur when microparticles penetrate the entireepidermis and become lodged in the underlying dermis.

[0108] This invention also relates to a method of detecting theappearance of blood within one or more microconduits, before bloodentering a microconduit moves out of the microconduit and then leavesthe tissue in which the microconduit(s) is formed. According to anembodiment, detection of blood within one or more micro conduits can beused to assess the size, particularly depth, of a rnicroconduit if thetissue is known to have blood vessels such as a capillary bed locatedaway from the tissue surface where microconduits are formed.

[0109] According to a preferred embodiment, a method for detection ofblood within one or more microconduits includes the use of optical meansthat employs reflected light which has spectral properties differentthan reflected light from the tissue in which the microconduit isformed. For example, measurement of the ratio of reflected red light toreflected blue light can give an indication of the entry of blood into amicroconduit.

[0110] According to an embodiment, for detection of blood within one ormore microconduits it is preferred to use image analysis that candistinguish a microconduit from the tissue in which a microconduit isformed. If a mask with microholes (FIG. 1) is used, then the edges ofthe microholes can be distinguished and used to identify the region inwhich microconduits are being formed (or were formed). Additionally, theedges of one or more microholes can be purposefully marked with distinctdyes so as to enhance the location of a region of interest (the areawithin a microhole).

[0111] If a collimated microparticle beam (FIG. 2) is used to form oneor more microconduits, the image analysis means can be directed to thegeneral tissue surface region. By using a previously calibrated beamsize, the image analysis means can be constrained to look for localizedareas corresponding to the beam size.

[0112] In an embodiment, a relatively inexpensive video camera with animage acquisition time in the range of between about one tenth (0.1) andabout two (2) seconds can be used if the microconduit formation time isin the range of between about one (1) and about twenty (20) seconds, asthis controls the microconduit depth to about ten percent. According toan embodiment, if a different depth resolution is desired, the ratio ofimage acquisition time to microconduit formation time can be chosendifferently. The camera's spatial resolution need be only sufficient toresolve the localized area of a microconduit, so that the appearance ofblood within the localized area can be detected. According to anembodiment, detection of blood within a microconduit by a video cameracan be used with any method of microconduit or skin opening formationthat is compatible with viewing of the microconduit or skin opening by avideo camera.

[0113] In one embodiment, the process of forming one or moremicroconduits by microparticle impingement can be followed by theadditional step of transporting one or more therapeutic molecules orions through one or more microconduits to achieve drug delivery totissue, including skin, for example.

[0114] In one embodiment, transdermal delivery of therapeutic agents(e.g. drugs such as insulin and genetic material such as DNA) isaccomplished by forming a microconduit according to an embodiment of theinvention, and then directing the therapeutic agent into themicroconduit, thereby delivering the therapeutic agent through the skinto the tissue. In another embodiment, transdermal extraction of analytesis accomplished by forming a microconduit according to the invention andthen removing the analyte from the tissue and through the microconduit,thereby removing the analyte from the tissue and through themicroconduit. In one embodiment, the analyte is removed by sampling. Forexample, the analyte blood is removed by allowing the blood to flow outof a microconduit onto a collection sheet or plate. In anotherembodiment, an analyte such as interstitial fluid is removed by using,for example, a pipet to reduce pressure over the microconduit.

[0115] Many other examples of drugs and genetic material are well known,including drugs such as lidocaine and other anaesthetics, heparin, low,erythropoietin, growth hormone, steroids, various peptides, and geneticmaterial such as large DNA segments, RNA, small antisenseoligonucleotides, and immunological material generally, includingvaccines and adjuvants.

[0116] Transdermal delivery of therapeutic agents through a microconduitaccording to an embodiment is important for a number of reasons,including the fact that often the intact stratum corneum preventstherapeutically significant rates of molecular and ionic transport.Microconduits according to an embodiment allow sterically unhinderedmovement of molecules and ions through the stratum corneum. The movementof molecules and ions through a microconduit according to an embodimentmay take place through diffusion, electrophoresis, or convection flowdriven by hydrostatic pressure differences, and time varying pressuredifferences including ultrasound produced and osmotic pressuredifferences. This includes iontophoresis which can involve bothelectrophoresis and electro-osmosis. In one embodiment, a direct currentvoltage is applied to a microconduit to produce iontophoresis. In aparticular embodiment, the direct current voltage applied to themicroconduit is pulsed.

[0117] According to an embodiment, molecular and ionic movement throughone or more microconduits with varying degrees of control can beachieved by using different amounts or concentrations of the moleculesand ions supplied. According to an embodiment, molecular and ionicdiffusion can be controlled by controlling the supply concentration ofthe molecules and ions, controlling the solution (usually based onphysiologic saline) used, and establishing or measuring the temperature,and then controlling the time that the supply solution is in contactwith one or more microconduits. One embodiment of the invention utilizesdiffusion of a therapeutic agent in a suitable pharmaceutical carrier,such as a biocompatible, non-toxic liquid, through microconduits toachieve transdermal drug delivery. In another embodiment, in order toachieve sustained release of the therapeutic agent, a therapeutic agentis supplied in a hydrogel, polymer, or molecular matrix, rather than ina liquid solution.

[0118] According to an embodiment, the invention relates to a method forforming one or more microconduits that allow desired molecular and ionictransport while substantially preventing the entry of infectious agentssuch as virus particles, bacteria and yeast. This exclusion ofinfectious agents is based on the formation of a molecular matrix withinthe microconduits. The openings within the matrix are generally smallenough to block infectious agents or severely hinder the entrance of theinfectious agents into the skin. A disadvantage of a molecular matrix isthat it decreases movement of molecules and ions by convection flowbecause the characteristic size of the openings within the molecularmatrix are small. However, movement of molecules and ions by bothdiffusion, electrophoresis, and electro-osmosis, can still be achievedwhen a molecular matrix is used according to an embodiment.

[0119] Aqueous gels that are biocompatible and can exclude infectiousagents while admitting ions and molecules are a suitable type ofmolecular matrix. Examples of gels are agarose, agar and carrageen.Polymer matrix gels can be positioned in contact with the skin by meansof a pressure-sensitive adhesive with a rate controlling membrane orlayer for the delivery of active molecules.

[0120] Another suitable gel is calcium alginate, which is gelled byexposing sodium alginate to an elevated calcium (Ca²⁺) concentration.Thus, if Ca²⁺ concentration is used to alter the recovery of amicroconduit, the high concentration that tends to maintain atransdermal microconduit open is generally compatible with the use of ahigh Ca²⁺ concentration to maintain a calcium alginate gel within amicroconduit. Conversely, decreasing the Ca²⁺ concentration near orwithin a microconduit tends to both encourage tissue recovery that willseal off a microconduit and simultaneously tends to dissolve the calciumalginate gel.

[0121] In addition to gels such as agarose, agar, carrageen and alginatewhich are obtained from natural sources, biocompatible polymer matricesobtained by cross-linking synthetic polymers can also be used. Thisincludes polymers, which have been described for use in controlledrelease of drugs. This includes poly(L-lactic acid), poly(DL-Lacticacid) and copoly(lactic/glycolic acid). It is well known that thepolymer matrix properties can be controlled by altering the polymermolecular weight or copolymer ratio (see Miyajima et al. “Effect ofpolymer/basic drug interaction on the two-stage diffusion-controlledrelease from a poly(L-lactic acid) matrix,” J. Controlled Rel.61:295-304, 1999). As another example, the transport of proteins out ofa hyaluronate matrix was relatively slow for a flly esterfied matrix butmore rapid for a less esterfied hyaluronate matrix (see Simon et al.“Mechanisms Controlling Diffusion and Release of Model Proteins Throughand From Partially Esterified Hyaluronic Acid Membranes” J. ControlledRel. 61:267-279, 1999). See also G. D. Prestwich et al. “ControlledChemical Modification of Hyaluronic Acid: Synthesis, Applications, andBiodegreadation of Hydrazide Derivatives” J. Control. Release 53:93-103,1998.

[0122] A variety of polymeric and bioerodable preparations suitable forimplantation and subsequent release of bioactive material have beendescribed in the scientific literature (see, for example, the review, R.Langer “Drug Delivery and Targeting” Nature 392:55-S10, 1998).

[0123] The invention also relates to a delivery method and apparatus,such that immunizing material can be effectively introduced into thetissue near dendritic cells, and other cells such as keratinocytes, andthen, as a further step, delivery into the dendritic cells,keratinocytes, and any other target cells within the skin.

[0124] According to an embodiment, the process of forming one or moremicroconduits is followed by the additional step of transportingimmunizing material into the tissue. Cutaneous immunization in whichimmunizing material is delivered to dendritic cells within the skin isof particular interest. Thus, formation of transdermal microconduitsthat fully penetrate the stratum corneum can be followed by transport ofimmunizing material into the epidermis.

[0125] This process includes the transport or delivery of nucleic acidssuch as DNA into the skin for the purpose of cutaneous immunization.According to an embodiment, a solution containing nucleic acids isapplied to the skin surface into which microconduits have been formed,and diffusion, electrophoresis or convection are used to transportnucleic acid molecules through one or more microconduits into the skintissue.

[0126] One or more nucleic acid molecules can also be transportedthrough microconduits into skin tissue for the purpose of gene therapy.

[0127] In yet another embodiment, the method comprises the further stepof creating an electric field so as to cause molecular transport withsignificant molecular transport component parallel to the stratumcorneum but within the epidermis. That is, stratum corneum-penetratingmicroconduits made by any suitable embodiment of the method can be usedto transport molecules into skin tissue; and suitable skin surfaceelectrodes such as ring-shaped electrodes around one or more suchtransdermal microconduits can be used to create an electric field with asignificant electric field component that is parallel to the outer skinsurface (see FIG. 3). An electric field with a significant componentparallel to the skin's surface thus causes lateral molecular transportwithin the epidermis by electrophoresis (electrical drift) and/or byelectro-osmosis. Such lateral molecular transport is useful to deliverdrugs and genetic material to skin cells away from the microconduitswhich are used to transport molecules across the stratum corneum. Suchlateral electrical transport is in addition to lateral diffusion, whichoccurs within the tissue below the stratum corneum or below a nail.

[0128] In another embodiment, additional control with a generally fasterresponse time is achieved by applying an electrical current or voltageacross the tissue. In one embodiment, in the absence of a geometricallyfixed, charged molecular matrix within microconduits, application of acurrent or voltage will predominantly produce molecular and ionictransport by electrophoresis, sometimes also called electrical drift. Inthe case of skin, transdermal drug delivery through at least onemicroconduit is thereby achieved by electrical drift (electrophoresis)as the transport mechanism. This can be achieved, for example, byproviding an electrode near a microconduit, and another, more distantelectrode at another site on the body. Alternatively, a tissue surfacering electrode can be placed with the microconduit opening within thering.

[0129] According to an embodiment, once microconduits have been formed,therapeutic agents such as DNA or other vaccine material, for example,are provided at the outer surface of the stratum corneum of the skin,and directed into the openings of the microconduits. Then, according toan embodiment, one or more transport driving forces such aselectrophoresis are applied, such that the therapeutic material orvaccine material is moved through one or more stratum corneum openingsand thereby transported into the epidermis.

[0130] The therapeutic or vaccine material is transported substantiallyparallel to the outer surface of the stratum corneum, and may thereforeencounter more immunizing cells than are located immediately adjacentthe openings through the stratum corneum. In addition toelectrophoresis, according to an embodiment, other suitable lateraltransport driving forces include use of electro-osmosis, hydrostaticpressure gradients, sonic and ultrasonic fields, osmotic pressuregradients and concentration gradients.

[0131]FIG. 3 is a schematic representation of an electrophoretic methodand apparatus, according to an embodiment, used to transport moleculesor ions through a microconduit and into the skin and parallel to a majorplane of the region of skin surface. In one embodiment, one type ofelectrode is positioned near, preferably substantially surrounding, anopening in a tissue such as the skin. In FIG. 3, the case of skin isshown, with a ring-shaped electrode 30 placed on the skin andsurrounding a microconduit, represented by a dashed line indicating theoutline of a tapered opening which comprises the microconduit. In theembodiment represented, the microconduit is wider at the outer surfaceof the skin, penetrates the stratum corneum completely, and terminateswithin the epidermis. The two cross-hatched vertical rectangles 34indicate the cross-section of an electrically insulating short tube,within which is an electrically conducting aqueous electrolyte solution(not shown). A central, “inner electrode” 32 is located in a reservoir(details not shown) above this tube, with this electrode shown withnegative polarity, suitable for electrophoretic transport of negativelycharged molecules from the reservoir through the microconduit andthereby into the epidermis, where the field lines spread out. Some fieldlines penetrate deeply through the epidermis and into the subcutaneoustissue before returning to the outer, ring-shaped electrode 30, hereshown with positive polarity. However, a significant number of fieldlines remain within the stratum corneum, and result in significant“lateral transport” of small ions and of charged molecules. This resultsin delivery of ions and molecules to regions of the skin away from themicroconduit or other skin opening. This is particularly useful fordelivery of DNA to the vicinity of cells within the epidermis that aresomewhat distant from the microconduit or other stratum corneumopenings.

[0132] In an embodiment of the invention, because of the size of themicroconduits, liquid convection flow operates naturally to movemolecules through a microconduit. Convection flow is important if nomolecular matrix has been formed within the microconduit. In oneembodiment, water soluble molecules are delivered by providing a drivingforce for convection through one or more microconduits. According to anembodiment, a pressure difference or pressure gradient is utilized todrive flow. For example, a pressure difference can be formed by applyingan increased pressure with respect to the pressure within the tissue, atthe terminus (entrance or opening) of a microconduit. If the surfaceopening of a microconduit is adjacent to a reservoir with adrug-containing solution of the molecule to be delivered, thenincreasing the pressure within the reservoir creates a pressuredifference along the microconduit, and flow results. This is analogousto applying pressure to the solution within the barrel (reservoir) of asyringe: Increasing the barrel pressure drives flow through the needle(analogous to a microconduit) into a tissue. Thus, if transdermal drugdelivery by convection through one or more microconduits is desired,flow can be established, according to an embodiment, by elevating thepressure in a drug reservoir that is held against the skin at the sitesof one or more trans-stratum corneum microconduits. For example, in aparticular embodiment, the drug or therapeutic agent directed into theopening is in a column, for example, a pipet or capillary tube, and thecolumn is sealed to the tissue around the microconduit. Next, pressureis directly applied to the microconduit, for example, by squeezing arubber bulb attached to one end of the column or pipet, thereby forcingthe therapeutic agent included in the column or pipet into themicroconduit. The therapeutic agent is absorbed by the tissuesurrounding the microconduit. According to another embodiment,convection through microconduits can also be established by using anosmotic pressure difference, a time varying pressure difference such asultrasound, and electro-osmosis.

[0133] According to yet another embodiment, pressure can also be used toforce a deformable drug-containing hydrogel from a reservoir or supplyinto one or more microconduits. When inserted into a microconduitaccording to an embodiment, the hydrogel can provide slow, controlledrelease of drug into the epidermis, or into deeper tissues if themicroconduit penetrates beyond the epidermis. Generally, moleculesreleased into the epidermis migrate so as to enter blood capillaries.Other types of slow release entities of small size can also beintroduced through microconduits.

[0134] According to an embodiment, molecular and ionic transport throughone or more microconduits in the outward direction from the tissue canbe used to acquire small samples of interstitial fluid, or a combinationof interstitial fluid and intracellular fluid if cells are permeabilizedor lysed, or blood if the microconduit extends sufficiently deep andaccesses one or more capillaries or blood vessels within the tissue.Samples acquired by a method according to an embodiment can be presentedto sensors or other measurement means located outside the body, and usedfor measurement or sensing of chemical analytes within the extractedfluid. Different types of sensors and assay systems which are suitablefor sensing or measurement of the extracted sample have been developedby others and are well known in the art.

[0135] Transdermal extraction of small fluid samples for transdermalmeasurement is an important embodiment of the invention, and isaccomplished by forming one or more stratum corneum-spannmgmicroconduits through which a sample is transported for the purpose ofcarrying out a transdermal analyte measurement.

[0136] The invention also relates to a general process in which one ormore analytes are measured while within a microconduit, rather thantransporting the analyte out of the tissue to be measured by an externalsensor or measurement means. For example, according to an embodiment, ifa microconduit is formed in skeletal muscle, cardiac muscle, bloodvessel wall or the liver, an optical measurement performed on fluidwithin a microconduit is used. This avoids problems associated withhandling of very small samples, such as problems associated withdilution or contamination. Suitably small sensors can be inserted into amicroconduit, or, preferably, optical measurement means such asnear-infrared Raman Spectroscopy (see, for example A. J. Berger et al.“Feasibility of Measuring Blood Glucose Concentration by Near-InfraredRaman Spectroscopy,” Spectochim. Acta 53:2887-292, 1997; T-W. Koo etal., “Reagentless Blood Analysis by Near-Infrared Raman Spectroscopy,”Diabetes Tech. Therapeut., 1:153-157, 1999; A. J. Berger et al.“Multicomponent Blood Analysis by Near-Infrared Raman Spectroscopy,”Appl. Optics 38: 2916-2926, 1999) may be used.

[0137] According to an embodiment, to make an analyte measurement withinone or more skin microconduits, the analyte can enter a microconduitfrom the tissue into which the microconduit penetrates, e.g., epidermaltissue. If the microconduit penetrates into a blood capillary bed andresults in blood entering one or more microconduits, then the bloodlevel or concentration of one or more analytes can be measured withouttransporting analyte through the microconduit out of the body. Instead,one or more analytes can be measured with the blood remaining within thebody by virtue of the fact that the microconduit is within the body.This is particularly desirable if an optical measurement method is used.Artifacts and errors associated with nonblood tissue can be greatlyreduced or essentially eliminated, because the blood is in full view ofthe optical measurement means such as reflectance spectroscopy,including near infrared Raman reflectance spectroscopy. An advantage ofmaking a measurement within the microconduit according to an embodiment,is that exposure of healthcare workers to biohazards such as HIV orhepatitis virus is greatly reduced.

[0138] If natural or stimulated processes cause one or moremicroconduits to become significantly closed, such that transport ofdesired ions or molecules is significantly sterically hindered, thensuch partially recovered microconduits can be reopened to achieve usefullevels of ion or molecule transport by application of pressure. Pressurecan be essentially held steady, such as for example, by applying suctionto open up a crust or initial formation of protective layer at the siteof a microconduit. Alternatively, pressure can be appliedintermittently, including in an oscillatory fashion. This includes theuse of ultrasound.

[0139] In some cases a transdermal microconduit will become fully orpartially blocked by lipids secreted from secretory granules that areinvolved in the skin's response to other types of stimulation. In thiscase, if the electrical resistance at the site of the microconduitbecomes large due to blockage or coverage with lipids, one or moreelectrical pulses can be applied to form aqueous pathways through thelipid layers. The use of such electrical pulsing can also be combinedwith application of pressure.

[0140] This invention can also involve the additional step of providinga stimulus that causes cells to take up molecules such as therapeuticmolecules and ions that have been laterally transported by an electricfield with such a parallel (to the tissue surface) component. Stimulisuitable for causing such cell uptake within the epidermis includeultrasound, heating and additional electric field pulses which causeelectroporation. An example of a cell that can be stimulated to take upa therapeutic agent is a dendritic cell. Examples of therapeuticmolecules or ions that can be taken up by a cell are DNA andanti-neoplastic drugs. Finally, one or more electrical pulses areapplied, such that at least one immunizing cell within the skin iselectroporated such that vaccine material is delivered into at least oneimmunizing cell within the skin.

[0141] In yet another embodiment, it is possible to practice thisinvention without the step of providing lateral transport, in this caserelying on delivery of vaccine material to the immunizing cells close tothe stratum corneum openings.

[0142] The invention also relates to a method of forming microconduitsthrough nail tissue.

EXEMPLIFICATION EXAMPLE 1

[0143] This is one of the first experiments carried out.

[0144] Materials and Methods: An S.S. White Airabrasive unit (S.S. WhiteTechnologies, 151 Old New Brunswick Rd., Piscataway, N.J.), including apressure chamber to hold an abrasive powder, was rebuilt. The pressurechamber was fitted with necessary valves, pressure regulators, and ahand piece on which a variety of nozzle sizes can be screwed; thepressure chamber was seated on top of a shaker. Using about 80 psinitrogen pressure, microparticles were used to scission the stratumcorneum of Subject A for about 12-15 seconds with the 500 μm diameternozzle held stationary, and approximately {fraction (1/16)} inch awayfrom his stratum corneum; no micromask was used. The microparticlescomprised alumina or aluminum oxide abrasive powder, similar to what hasbeen used in commercial units for facial peels. The microparticles weresharp, with size generally ranging between about 15 μm and about 20 μm,with some 30 μm outliers.

[0145] Results: Subject A reported a very slight sensation, andmicroscopic examination revealed a hole of indeterminate depth in hisstratum corneum.

[0146] Method and Materials: The experiment was repeated, impinging theabrasive stream on a 250 μm thick, plastic micromask having 800 μmdiameter holes. The micromask was taped to Subject B's left forearm onthe palm side, and he waved the nozzle over pairs of holes in themicromask for time intervals of 10, 30 and 100 seconds at a distance ofabout ⅛ inch or 3000 μm between the nozzle and the mask.

[0147] Results: Nothing was visible on Subject B's stratum corneum inthe 10 second case, but he could see capillaries in the bottoms of themicroconduits made during the 30 and 100 second exposures. Furthermore,those microconduits began bleeding. The microconduits were slightlylarger than the mask openings (microholes), and there may have been aswelling reaction up his forearm following the microparticleimpingement. Subject B reported sensing a very slight “pricking” feelingonly during the microscissioning process for the 30 and 100 secondexperiments.

[0148] The area along the forearm of Subject B where the holes were madeshowed signs of swelling. Later, a conclusion was reached that theswelling and inflamation were probably a sterile cellulitis, caused bythe very aggressive use of 100 second exposure to the air jet with themicroparticles, and that air was injected into the deeper tissue. Havingthereby learned about “air injection injury,” it was concluded that thiscould be easily avoided by using a combination of shorter exposures andsmaller air pressure.

[0149] The white light images of FIGS. 4A and 4B are labeled with arrowsmarked 1 through 6. In FIGS. 4A and 4B, microconduits 1 and 2 are theresult of the 100 second microparticle impingement; microconduits 3 and4 of 30 seconds; and 5 and 6 of 10 seconds. In the black and whiterendition of the original color photomicrograph, the blood drops at thesites of the two microconduits made by the 100 second exposure are darkgray, the smaller blood drops associated with the 30 second exposure arealso gray, and the microconduits at sites 5 and 6 are not visible. It isbelieved that microconduits 1 through 4 fully penetrated the stratumcorneum and epidermis, reaching blood capillaries, with blood flowingout due to normal blood pressure. However, the microconduits at sites 5and 6 are believed to have not fully penetrated the epidermis (novisible blood; no visible mark or sign within the first few hours afterformation of the microconduits.)

EXAMPLE 2

[0150] The experiment relates to the design of a mask holder toeliminate gaps between the mask and surface undergoing scission. SubjectA recognized the need to have a picture-taking capability which wouldcope with the normal shake (tremor) of the human forearm, since thetremor is magnified in a microscope. He therefore obtained a framegrabber TV camera package and worked with a zoom microscope and laptopcomputer. Although not perfect, it makes much better pictures (via acomputer and color laser printer) than were possible with the colorinstant camera used earlier.

[0151] Methods and materials: With this imaging improvement, Subject Abegan experiments with a micromask and microparticles. It was decidedthat holes on the order of 100 μm diameter on 500 μm center-to-centerdistances would be a reasonable place to start. Using 25 μm thickpolyimide (PI) mask material, Subject A impinged microparticles to formfour 100 μm holes. Holding this with a bandage that had a clearance holeof 1000 μm diameter, Subject A impinged microparticles onto themicromask for various times, up to 60 seconds with a nozzle having a0.016 inch (400μm) diameter opening.

[0152] Results: There was little or no penetration of Subject A'sstratum corneum, although dust that was visible on Subject A's skinindicated that aluminum oxide microparticles clearly got through. Twothings were noted. First, since the microparticles were in powder form,with grain sizes measuring 20 to 30 μm, it is likely that themicroparticles were too big to get through the microholes in themicromask. Second, the thin PI mask, held against Subject A's skin witha bandage, did not touch the skin surface. Instead, Subject A's stratumcorneum appeared to be pushed away by the bandage. This means that evenif the microparticles got through, they did not hit the skin, but spreadout, and the incoming air blast could actually “balloon” the stratumcorneum pocket (region) under the PI mask. This in turn could produce astagnating air bubble that would exclude most of the fresh incoming airand abrasive microparticles. It was therefore concluded that the maskneeded to be tight against the stratum corneum, and much thinner thanthe hole diameter to permit air and microparticle exchange. Subject Athen made a fixture, which acted somewhat like a drum head holder, withthe PI mask stretched across it and held with double-sided sticky tape.The mask “drum head” surface protruded about 0.250 inch above theholding handle. With the micromask pressed into Subject A's stratumcorneum, there appeared to be no space between the PI mask and stratumcorneum. Subject A then used a 60 second sandblast at 80 psi through a0.016 inch (400μm) nozzle on a one hole (microhole) mask.

[0153] The above procedure opened up the hole in the micromask to about5001im rather than 100μm. When the mask was used for microconduitformation, a clearly-defined hole (microconduit) was produced in SubjectA's stratum corneum with no sensation. The microconduit looked moist atits bottom, and when squeezed slightly with tweezers, it bled. It wasapproximately 250 μm to 300 μm in diameter. It is likely that there weretwo effects. One is that, as with any open hole type of mask or stencil,the material coming through has no effect near the edges of the openingbecause of air stagnation (friction effect) and possible piling up ofmaterial in the comer formed by the mask, and the stratum corneum. Thismaterial “self masks” the stratum corneum near the edges. Second, themicroholes must be some factor bigger than the microparticles goingthrough them, or a dynamic blocking effect will occur wherein onemicroparticle nearly blocks or knocks an incoming microparticle away.Clearly, there was a 100 μm per side difference between the PI mask hole(500 μm) and the resulting hole in Subject A's stratum corneum (3001μm).

EXAMPLE 3

[0154] The planned experiment was to test a confocal microscope, used tomeasure microconduits in vivo. The microscope, designed to look downthrough skin, uses a laser emitting 680 nanometer wavelength light asthe light source, coupled with a TV framegrabber and precise verticalobjective lens positioning. The wavelength of the illumination is suchthat materials with the refractive index of water are essentially clearor invisible, and materials with a refractive index away from thatnumber are visible. For this reason the skin (mostly comprised of water)becomes essentially transparent, and the image is that of the materialsthat are not water, such as organics and inorganics at boundaries in thestratum corneum, and other materials located deeper in the skin tissue.Thus, with the confocal microscope and laser light source, one can viewtissue changes that are several hundred micrometers deep in the skin.

[0155] Materials and Methods: The plan was that Subject B and Subject Awould each have an opening scissioned in their stratum corneum, to somedepth shy of blood, and to use different microparticle impingement timesin making the microconduits. More specifically, the plan was to form onemicroconduit per person, covered with a deionized water (DI) soakedbandage, then view the microconduits as soon as possible under aconfocal microscope. A new, 75 μm thick PI mask with a 160 μm diameterhole was used. An electrical resistance meter, also referred to as an“impedance meter,” was connected to two separate but adjacent conductivepoles that were in direct contact with the skin. Both tests started withan ethanol rinse, followed by a deionized water wipe of the area to beimpinged with microparticles. An EKG electrode was affixed to thesubject between the mask fixture and the heel of the palm. Theresistance of the skin between the EKG electrode and the area to beimpinged was measured by means of an electrical probe immersed inconductive saline that covered the mask/microhole and the EKG electrodebefore, during and after impingement. The results are tabulated below.SUBJECT Subject A Subject B Resistance Resistance DRY PI = Holder to EKG218 kohm 1.2 Megohm DI-I-suction-out 2.5 kohm 0.8-1.6 Megohm Saline 16kohm (shunt) Blow off 1.4 Megohm Saline 170 kohm 145 kohm Suction-outsaline 2.3 Megohm 600 kohm DI rinse + blowoff 5 sec. Microscission 400kohm 3 Megohms SC @ 60 psi Saline 160 kohm (possible shunt) 5 kohm(shunt) Rinse; suction; blowoff 2.9 Megohm 2.6 Megohm Saline 216-218kohm 20 kohm DI rinse 2.5 Megohm 1 Megohm-(No blood visible) 5 sec.Microscission 2.5 Megohm SC @ 60 psi Saline 30 kohm Double DI rinse,suction .4 to 1.2 Megohm Subject A - (Blood visible)

[0156] The sites on each wrist were marked with small, inked-in lines.The maskholder was cleaned with ethanol between Subject A and Subject Btests. The sites were covered with bandages soaked in DI water. Aconfocal microscope described above was used to examine Subject B's sitefirst. The time period between the test and the confocal examination wasapproximately one hour.

[0157] Results: The opening of the microconduit at the top of thestratum corneum was on the order of 100-125 μm. The depth was in therange of 130 μm. There was particulate debris lying on the surface andin the hole. It was difficult to determine the depth precisely, becauseillumination in the hole was poor. This was due in part to the backreflection being suppressed by the non-planar surfaces at the bottom ofthe hole. At depth, back reflection was further suppressed by the formfactor (depth to diameter ratio) causing increased collimation whichreduced the back reflection even more. However, the confocal microscopeworked very well, showing the cross sections of the hole, as made by themicroscissioning process, at selected depths. In general, themicroconduits tapered to half diameter at their bottoms, in the outlineof a blunt-bottomed carrot.

[0158] FIGS. 5A-5K include Subject B forearm microconduit confocalimages at different depths (indicated in μm from the appropriate skinsurface), starting at zero, and going to a depth of 170 μm. Note thatdeeper than 79 μm, there is less evidence of a microconduit.

[0159] Three percent (3%) lidocaine was applied on the Subject B site.No discernible anaesthetic effects were observed.

[0160] The examination of the Subject A's stratum corneum showed similareffects. The diameter at the stratum corneum was also in the 100-25 μmrange. The depth was approximately 175 μm. The blood clot in the SubjectA microhole was invisible with the confocal microscope. There appearedto be debris at the bottom of this hole also, as small (20/30 μm)spherical objects could be seen.

[0161] FIGS. 6A-6I include Subject A forearm microconduit confocalimages at different depths (indicated in μm from the approximate skinsurface).

EXAMPLE 4

[0162] A fungus is sometimes found to live in the interface betweennails (in particular, toe nails), and the underlying living tissue. Suchinfections tend to be chronic, since it is difficult to treat topicallydue to its protected location. An established treatment in serious casesis removal of the nail(s) to reach the fungus. Therefore, an experimentin making through-nail microconduits, to permit application of topicalantifungal chemicals, was performed.

[0163] Materials and Methods: Microparticle impingement was achievedusing a “particle generator”, which was a custom modified S.S. WhiteAirbrasive Unit, Model K, Series II (Piscataway, N.J.). This apparatusoccupies a cubical volume of approximately 14 inches on a side, andcontains a solenoid-driven table, onto which is bolted a pressurizedreservoir with a spring-loaded top, through which the microparticles canbe loaded into the reservoir. A tight fitting, 1.5 inch diameter×0.5inch plate, with a hollowed-out region, creating a circular particle/gasmixing chamber, was used. Carrier gas was admitted through a hole in thechamber wall, and exiting through a second hole in the wall,approximately 120 degrees away from the input. The thin top of thischamber served as the bottom of the reservoir, and contained a number ofsmall holes through which (microparticles) could fall into the swirlinggas stream of the chamber below. The particle chamber and mixing chamberwere at the same pressure. From the mixing chamber the particles/gasflow proceeded through a flexible plastic tube, which terminated in thenozzle holder. Various sized tungsten nozzles can be screwed onto thisnozzle holder, and act to direct and size the particle stream. The unitalso contained a power supply and meter to change the shake tableamplitude, which increased or decreased the flow of the particles intothe gas stream. Also there was a pressure gauge and pressure regulatorto monitor and set the gas pressure.

[0164] It should be noted that the air pressure used in this example (20p.s.i) was much lower than the pressure used for industrial applicationswith this machine. Further, the amount of particles in the flux ofparticles was much less than the amount of particles used in otherapplications. It should also be noted that having very few particlesgoing into the flux works better than having many particles going intothe flux. Otherwise, in forming a microconduit, a high concentration ofparticles in the flux will tend to pile up in the tissue, rather thanmicroscission and remove tissue.

[0165] Viewing and measuring the microconduits was accomplished with aBausch and Lomb Stereozoom 7 microscope with a variable magnification(2×-7×). A measuring eyepiece was used to check planar dimensions. APanasonic CCD television camera with a 0.75×-3× lens connected to a 15inch color monitor was used to view regions of test material, skin ornail before and after microconduit formation protocols. The monitor wascalibrated against a vernier caliper projection (image), with marks handdrawn onto the monitor screen. This could also be connected to acomputer having a frame grabber (Visionx), which allows the image to beviewed and recorded on a floppy disk for making color photomicrographs.

[0166] Microparticle impingement was carried out using the 0.011 inchnozzle at 20 psi and 70v flow (setting on the S.S. White machine),spaced approximately 0.030 inch between nozzle and nail surface. The0.125 inch or so of overhang edge of the nail on Subject A's left littlefinger was used.

[0167] Results: That area of the nail, 0.020 inch (500μm) thick, wasscissioned through in 15-20 seconds (best estimate is 16 seconds).Because no mask was used, the microconduit was larger on the nozzle sideand tapered to 0.010 inch diameter on the far side of the nail. Themicroconduit opened quickly, possibly because the nail is much harder,less “bouncy” than the stratum corneum, and the nozzle was half as faraway from its surface.

[0168] The experiment was repeated on another area, but this time,Subject A scissioned part way through the nail, and put some black inkin the resulting “cup” (depression or incomplete microconduit) to see ifit would soak in, or diffuse beyond, the dimensions of the “cup.” Itdidn't—there didn't seem to be any lateral diffusion of the ink at all,which was surprising, as nails are brick-wall like, in the same fashionas the skin's stratum corneum, only much thicker.

[0169] Although not then known to Subject A, it would be useful to knowwhat chemical indicators might exist to determine how close to thenail's bed the microconduit has approached. Clearly, the microconduitformation rate difference between stratum corneum and nail (somewherebetween a ratio of 7 and 15 to 1) means that the microconduit formationmight well “self-limit” (stop) when the softer substrate (underlyingtissue) is reached. It acts like the skin that isn't beneath the nail.To know when to stop impingement, one could rely on the “sensation”indicator (a small prickling feeling or the appearance of blood), butthere may be other indicators possible. A question raised was, how closedoes a microconduit have to extend through the nail to deliver drug foreffective treatment of the fungus?

[0170] It was recognized that it would be useful to devise a fixture tohold the nozzle a fixed distance above the nail. It is important tostabilize the nozzle to nail spacing, and also tie the nozzle to thenail's motion. Trying to place the finger on a flat surface under thefixed nozzle relies on the spongy finger pad, which clearly leads to toomuch motion over a 10-20 second time interval. This fixture probablyshould be strapped onto the nail, like a saddle on a horse, with thenozzle held rigidly to this frame.

[0171] There are many potential applications for nail microconduits, inaddition to treating fungus. For example, identification numbers couldbe formed, using standard arabic numbers, bar codes, digital codes, orthe like. The removal of less than full depth would allow one to “write”on the nail surface. This may also find cosmetic application, in whichpeople write the initials or names of friends and loved ones, perhapswith coloring (dye) added to the “etched” region. Within a few weeks,the outgrowth of the nail would carry such markings away, to be removedwith nail trimming.

[0172] For cosmetic purposes, a light scission by microparticleimpingement could improve the adherence of a faux nail to the real nail.Bas reliefs can be engraved in the nail surface, and dyed. One couldpossibly put in dye at various levels by partial excission, dying, thenscissioning a little deeper and dying a different color below, forexample.

EXAMPLE 5 Lidocaine 2

[0173] This experiment involved creating a transdermal microconduitfollowed by delivery of lidocaine to attempt achievement of localizedanesthesia.

[0174] Methods and Materials: A single microconduit anaestheticexperiment using fifty percent (50%) Lidocaine was conducted. Themodified S.S. White apparatus was used with a setting of 35.7 Volts, 25psi. With these settings, a microscission-through time of 18 seconds wasestablished in test material. The mask was a single 170 microhole in 3mil kapton. The preparation protocol included application of ethanol, aDI water soak, and drying. The initial resistances were Dry—open (opencircuit; very high resistance), Saline—230K (kohm), Suction out/DIrinse—open.

[0175] Microscission by microparticle impingement was carried out for 10seconds, which would give a microconduit depth that was clearly throughSubject A's stratum corneum, but would not exceed 50 -70 μm, based onthe recent data from the confocal microscopy experiments.

[0176] The resistance was Dry—open, Saline-11 kohm (a significantdecrease) and Suction out/DI rinse—open. Cotton swabs were soaked in theLidocaine solution, and one was placed on a control(non-microscissioned) stratum corneum control site, and the other on themicroconduit, which was clearly visible under the microscope. A swab washeld on the microconduit site for 2 minutes. A second swab was held onthe control site for 2.5 minutes. A new No.22 hypodermic needle was usedto prick probe to determine existence of sensation.

[0177] Results: A slight pressure on touching the stratum corneum withthe point produced sharp sensation. Using a scale divided into 32nds ofan inch (0.031 inch), Subject A pricked his stratum corneum at differentradial distances from the microconduit. His skin was numbed to the pointof no sensation for approximately 0.015 inch (half a space), and thensensation was clearly felt around the {fraction (1/32)} inch distanceand farther out. Subject A could not state that at the {fraction (2/32)}inch marker there was any different sensation level than untreatedstratum corneum (if it did, it was insignificant). Subject A thoughtthat the region around the microconduit became more numb 4 or 5 minutesafter the treatment. The numbness began to diminish after 9 minutes. Thetreated control site showed the same level of prick sensation as anontreated site.

EXAMPLE 6 Lidocaine

[0178] This experiment involved creating a transdermal microconduitfollowed by delivery of lidocaine with applied pressure to augmenttransdermal drug delivery, in this case for achievement of localizedanesthesia.

[0179] Subject A used a standard pin, which is slightly duller than thehypodermic needle, but can still be felt clearly, and doesn't damage thestratum corneum or outer epidermis as much as the sharp needle. 10 mgepinephrine added to a 20 ml saline +1 gram lidocaine (50% lidocaine)mixture was prepared.

[0180] Initially, when trying to calibrate the modified S.S. White“microparticle generator” before the experiment, the test microscissionrates in 2 mil kapton strips were extremely erratic, but then becamestable. Then, with a microscission time through 2 mil Kapton of 24seconds (33.46V), Subject A used the same 150 μm diameter single holemask on the new maskholder for the experiment below, with 8 secondimpingement time. Resistance Dry = 125 K kohm DI water = 1.5 Meg (Meg =Megohm) Suction out; blow out = 124 kohm Saline = 150 kohm SO (suctionout) = 213 kohm DI = 500 kohm Suction out = 115 kohm DI = 450 kohmSuction out, Blow out = 150 kohm

[0181] Scission by microparticle impingement was conducted for eight (8)additional seconds; at first obtaining a slight amount of clear fluid,and later a very slight amount of blood. DI = 105 kohm Suction, Blowout= 560 kohm DI = 800 kohm Suction, Blowout = 250 kohm Saline = 6.3 kohmSuction, Blowout = 300 kohm DI = 115 kohm Suction, Blowout = 360 kohmSaline = 6.5 kohm Suction, Blowout = 300 kohm DI = 500 kohm Suction,Blowout = 200 kohm

[0182] An attempt was made to increase the delivery of 50% Lidocaine+epinephrine (the “drug”) through the microconduit by applying pressureto the outer (skin surface) opening of the microconduit. The followingresults for the increase in numbed area are: Drug Water pressurePinprick numb radius 1 min. 30 inch  400 μm 2 min 30 inch  800 μm 3 min30 inch 1600 μm 6 min 30 inch 2800 μm

[0183] Results: First, there was no observable blanching of the stratumcorneum/epidermis around the microconduit site. Subject A tried the samepressure, the same drug (lidocaine-epinephrine) on another (control)site (no microconduit) to check possible changes, but could really seeno difference from the surrounding skin. But his skin pigmentation was“white”, so any blanching may have been difficult to discern. Also, it'spossible that the epinepherine didn't really reach enough capillaries,or that near-stratum corneum capillaries don't respond the same way asdeeper blood vessels.

[0184] Second, testing with the pinprick approach yielded the sameresults as before. However, Subject A continued pin pricking to findthat approximately 6 (six) minutes after the last injection(introduction through the microconduit), the radius of numbness was inthe vicinity of 2800 μm. This outer limit began shrinking after 10-12minutes. Thus, there seemed to be an additional anaesthetic spreadingeffect for about 6 minutes.

[0185] Third, Subject A tried electrical stimulation as a test of theextent of anesthesia. He used 200 volt, 2 ms, 30-40 mA pulsing (twopulses per site) after discovering the 6+ minute lateral spread effect.The site was numb to the electrical pulses within a 1400/1600 μm radius.Previously there was only slight sensation to the electrical pulses nearthe microconduit, much less than in non-anesthetized regions. Withpressure-driven delivery through the microconduit, however, these strongelectrical pulses weren't sensed, suggesting delivery was to a greaterdepth (and greater extent laterally) over the 6 minute period.

EXAMPLE 7 Glucose 1

[0186] This experiment involved creating a transdermal microconduit toproduce a blood sample that would be then used for a blood glucosemeasurement.

[0187] Methods and Materials: The experiment is intended to measureblood glucose, using a commercially available device that can measureglucose in a drop of blood. The measurement device is a “Lifescan” “FastTake” Glucose Sensor, with a readout contained in an oval caseapproximately 1.5 inches×2 inches by ⅜ inch thick. On one edge is a slotinto which is plugged that 0.031 inch×0.210 inch×1 inch sampling strip.This is plastic, has three electrical contacts on the plug in end, and asmall capillary, 0.1 inch×0.1 inch×0.010 inch, in which appear to be twoelectrodes, or gel areas, each of which are 0.1 in×0.05 in, one in frontof the other, so that the blood being pulled into this chamber bycapillary action must cover all of the first before beginning to coverthe second. Once the sample chamber is filled, the electronics takes 15seconds to display the glucose level (50-200 mg/dL).

[0188] Subject A carried out a 16 second microlocalized microparticleimpingement, using a mask with one 150 micrometer microhole, to form amicroconduit that should reach blood capillaries. Subject A squeezedaround the microconduit once in the x-x direction, and once in the y-ydirection, obtaining enough blood to fill the rectangularcapillary/measuring chamber for the measurement device.

[0189] Results: The instrument took 15 seconds to respond, and displayed97 mg/dL for Subject A's glucose level, which is a normal value. Thisprocedure was very simple and easy. There was little sensation.

EXAMPLE 8 Glucose 2 This experiment also involved creating a transdermalmicroconduit to provide a blood sample that would be used for a bloodglucose measurement.

[0190] This experiment again used a commercially available device (the“Lifescan” “Fast Take” Glucose Sensor) that measures glucose in a dropof blood.

[0191] A 10 second (microparticle impingement) exposure on the arm ofSubject A was conducted, to determine how much less exposure time wouldwork in measuring the blood glucose level.

[0192] Results: The electrical resistance after exposure was 50 kohm,and microscopic examination through the mask showed little redness, soSubject A did another 10 second exposure to make sure that blood vesselshad been reached. Blood was indeed obtained, and the electricalresistance was now 7.7 kohm. Blood flowed out. Subject A used a teststrip that came with the kit in the meter, but it's detector lookeddifferent. It had a detector area twice as long as the prior strip, andthere was no clear cover over it, so it couldn't draw out blood bycapillary action. Instead, a blood drop had to form, and be dropped inthe sample area of the test strip or the test strip laid face down init, rather than have the end edge dipped in and the blood suck up intothe capillary. There was enough blood to get a reading of 107 mg/dl. Aswith Example 7, the sensation detected was just slightly discernible.

EXAMPLE 9 Nail 1

[0193] This experiment involved measurements of the rate of microconduitdepth formation (“scission” rate).

[0194] Materials and Methods—Additional experiments use “cantileverclippings” from Subject A's toe nails. These toe nail specimens behavethe same way as finger nails, with nearly the same scission rate(0.00075 inch per second). To further determine the rate of microconduitdepth formation more toe nail clippings were used.

[0195] Microlocalized scission (microconduit formation) was carried outin experiments on Subject A's toe nail clippings, with the results shownbelow. TABLE Subject/experimenter: Subject A/Subject A Thickness atmicroconduit Time (sec) Scission rate (in/sec) TOE 0.015 inch (375 μm)15 0.001 0.025 inch (625 μm) 35 0.0007 0.030 inch (750 μm) 51 0.000580.042 inch (1,050 μm) 210 0.0002 FINGER 0.017 inch 17 0.001

[0196] Results: Based on previous experiments, there were no surprises.The decrease in scission rate (microconduit depth increase rate) withthickness was perhaps due to “self masking” of incoming microparticlescompeting with backbouncing microparticles, with this competitionincreasing with increasing form factor. Based on present understandingof the microconduit formation process, the major source of variabilityis due to the nozzle angle to the nail surface.

[0197] Some variations of the particle generator parameters wereconsidered next. The nozzle-to-nail surface space was always 0.030 inch.

[0198] 1. Vary pressure on a 0.016 inch (400 u) thick finger cantilever(flow setting=85 V, as in past tests) Pressure (psi) Time (sec) Scissionrate (in/sec) 15 34 0.00047 20 16 0.001 25 12 0.00133

[0199] 2. Vary pressure on a 0.016 inch thick finger cantilever (flowsetting 65V, changed from past tests) Pressure (psi) Time (sec) Scissionrate (in/sec) 15 25 0.00064 20 21 0.00076 25 15 11 0.00107

[0200] Again, with the exception of the 15 psi (lower flow result),there were no surprises. The low flow (65V setting) test was repeatedtwice, with nearly identical results. Perhaps it is worth exploring thelower flow, lower pressure regime, as well as higher pressures. Since,however, sensation appears not to be a problem, it might to bereasonable to increase the pressure to achieve faster microconduitgeneration into the nails. In both the above tests, the microconduitsmeasured 20/22 mils at the “top” (outer) surface and 15/16 mils at thebottom side.

[0201] All these tests were done with the nail clippings held in afixture with their “outer end” (outer surface) up, as if the hand wereheld vertically. The nozzle was located on a second fixture, withmicroparticle impingement horizontal. Both holders (nail specimen holderand nozzle holder) were mounted on x-y-z manipulators, with thetarotation to permit the nozzle to be positioned (by eye) perpendicular tothe nail's “outer” surface. This arrangement allowed one to observe theback (former inner surface) of the clipping while scissioning, to seewhen the through-microconduit (penetrating microconduit) first appeared.One could also shine a light at the nozzle side to make the penetrationprocess easier to observe. However, it turned out that the bestindicator was that of a microparticle pattern (“dust”) appearing on thefloor beneath the nozzle/nail arrangement, scattered by the N₂ (nitrogengas) stream coming through the nail, rather than bouncing back from theouter (front) surface.

[0202] In looking at one of the test fingernails, it was noticed thatthere was a good display of the kinds of microconduit centers one canachieve. The dimensions of Subject A's nails are, Little = 0.350 inch W(width) × 0.300 inch L (length) Middle = 0.450 inch W × 0.400 inch LThumb = 0.550 inch W × 0.400 inch L

[0203] All measurements are 90% of the maximum dimension. This meansthat if one can easily open microconduits on centers from 0.020 inch orgreater, it is possible to span nails with properties like those ofSubject A with anywhere from 5 to 20 microconduits of 0.015-0.017 inchmaximum diameter. If one wanted to form a full field of microconduits,it would require 25 to 360 microconduits, as an example.

[0204] In another application, one could think of a nozzle being drivenX-Y by stepping motors which interface with a computer programmed with asimplified image converting or numerical control machining program. Onecould sculpt a variety of images, numbers, pictures within a few minutesper nail. In the case of complex images, the computer program could alsomodulate the flow and/or pressure to produce depth variations whichwould enhance the “dimensionality” of the sculpture.

[0205] A check was done on high pressure N₂, at 30 PSI, 85V flowsetting, 0.030 inch nozzle-nail spacing, and the through-microconduittime was 9 second on the 0.016 inch thick part and 11 second on the0.018 inch thick part of that nail. This corresponds to 0.0018inch/second and 0.0016 inch/second.

EXAMPLE 10 Nail 5

[0206] This experiment involved measurements of electrical resistanceassociated with nail microconduits, and commentary related to antifungaltreatments.

[0207] Electrical resistance measurements have been used in anembodiment of the invention to partially characterize skinmicroconduits. At this point, it had been two weeks since the firstmicroconduit accessing blood was formed in the left hand ring fingernailof Subject A. However, we anticipated that there would nevertheless be alarge resistance decrease associated with the microconduit, becauseinsignificant nail repair was expected, and other protective layers thatform should have a much smaller electrical resistance than the nail.

[0208] Materials and Methods: The LCZ meter (a device for measuringelectrical resistance and impedance in skin or transdermalmicroconduits) parameters were the usual 1 V, 1KHz. All resistance testswere checked for greater than 5-10 Megohm “before test” readings. Atthese resistance levels, Subject A took special precautions to isolatehimself, from anything, especially shunting to the EKG (ECG,electrocardiograph) electrode, through his right hand that was holdingthe Tungsten (W) probe, a small, stiff electrically conducting wire thatcould be inserted into a microconduit. The tungsten wire was wrappedaround a 6 inch long plastic rod, which Subject A held in a glasshypodermic syringe while wearing a plastic glove. Subject A always kepthis left hand/arm elevated from the table during the tests. Results ofthe experiment are summarized in the following table. Results andAnalysis: Resistance 1.2-0.005 inch diameter Tungsten wires in Saline =2.2 kohm 2. EKG in saline, W wire in saline = 1.3-2 kohm 3. EKG on LHmiddle finger tip, EKG in Sal on = 11 kohm middle finger 4. EKG on LHmiddle finger tip, EKG on LH ring = 35 kohm finger tip 5. EKG on W wiretouching stratum corneum on = 1.5-15 Ring fing., Megohm 6. EKG on Wtouching DI drop on stratum corneum = >500 kohm on Ring fing. 7. EKG onW touching stratum corneum in DI drop on = 130 K-150 stratum corneum onRing fing. kohm 8. EKG on W touching Sal drop on Stratum corneum = 31kohm on Ring fing. 9. EKG on EKG on LH ring finger nail = 500-550 kohm10. EKG on W in Sal on LH ring finger nail = 500 K to > 10 Megohm 11.EKG on W in DI H20 on LH ring finger nail = 260 kohm 12. EKG on W inSal. Drop in large microconduit = 130 kohm to nail bed 13. EKG on W inSal drop in microconduits to nail = 120 kohm bed and near nail bed

[0209] From these measurements it is clear that the two microconduits inSubject A's LH (left hand) ring finger nail have led to a factor of 5reduction in the EKG-on-LH-nail-to-EKG on LH middle finger tipresistance (9, 12. & 13, above). A clean, “saline in small microconduitonly” measurement, could not be obtained because this microconduit wasclose to the large microconduit.

[0210] A review of relevant information about nails appears in, “TheNail”, Vol. 1, Physiology, Biochemistry, and Molecular Biology of theSkin, Second Ed., 1991 Lowell A. Goldsmith, Editor, the teachings ofwhich are incorporated herein by reference in its entirety. The term“nail” usually refers to the nail plate, which is a hard, clear towhitish, rectangle of tissue on the dorsum of the distal phalanx. Inorder to design an in vivo experiment to determine whether placingfungus-killing chemical compounds into one or more microconduits througha toe nail, some additional information may be needed. Clearly, however,a microconduit removes essentially all of the barrier function of thenail. With this in mind, a general review of the nail size and thicknesson the left side of Subject A was determined. Little Ring Middle IndexThumb Left Hand Length 0.300 inch 0.350 inch 0.350 inch 0.400 inch 0.425inch Width 0.350″ 0.450″ 0.475″ 0.375″ 0.550″ Thickness 0.016″ 0.017″0.018″ 0.021″ 0.026″ Left Foot Length 0.100 inch 0.225 inch 0.350 inch0.350 inch 0.550 inch Width 0.325″ 0.325″ 0.400″ 0.375″ 0.550″ Thickness0.025″ 0.025″ 0.039″ 0.036″ 0.053″

[0211] It is clear to see that the toe nails are thicker than fingernails by a factor of 1.5 to nearly 2. That means that toe microconduitswill be bigger than have been found previously by us for finger nailmicroconduits, where fingernails have been tested—mask or nomask—because the undercut in a 0.030 inch-thick nail will tend toproduce a microconduit approximately 0.020 inch diameter, minimum, evenif a mask with a 0.006 inch diameter is used. In addition, even with nomask, the microscission-through time would be in the range of a minuteor more per microconduit. The approach in the experiment was to carryout microconduit tests without a mask. Toe nail sizes are both biggerand smaller than fingernails, which will perhaps require severaldifferent sized nozzle holder “saddles.”

[0212] The question of whether to form many microconduits in a nail,located, for example, on centers twice the microconduit diameter, wasconsidered. An alternative, exemplary approach is to form a line ofmicroconduits across the nail, back towards the lunula. It is believedthat fungi only grow on dead material. For this reason, the under-nailfungal infestation is living on the nail only, not the nail bed, nor onthe nail-originating tissue known as the nail matrix, which is beneaththe skin, and under and behind the cuticle, and somewhere out into thevicinity of the lunula. With all this in mind one application of theinvention may involve a line of microconduits across the lunula edge ofthe nail, with the microconduits acting as reservoirs and “sweeping” thefungus out as the nail moves out.

[0213] Antifungals useful in an embodiment include non-prescriptiondrugs such as Tolnaftate including Tinactin(g, or Miconazole NitrateCream USP, 2% (E. Fougera & Co., Melville N.Y., 11747), Terbinafin, orprescription antifungal drugs such as Penlac®. The non-prescriptiondrugs are typically used as topical applications to kill and/or blockreplication of dernatophate fungi which cause “onychomycosis”, whichcauses ringworm and athlete's foot (tins pedis). With the presentexception of Penlac®, there is no approved topical drug for treatment ofnails, presumably due mainly to the inability of significant drug toreach the site of most of the fungus. The active ingredient ofTinactin(® is tolnaftate; Miconazole nitrate is itself the activeingredient of the second nonprescription cream-form medication.

[0214] Penlac® is used to attempt to kill the same fungus when it occursbetween the nail and nail bed. It is used in the form of a laquercontaining the antifungal chemical “ciclopirox”, typically provided insolution at 8% concentration. Loprox(is another antifungal preparation,which has the same antifungal agent (ciclopriox) except at a smaller(0.77%) concentration, which is available in cream, lotion or gelcombinations for treatment of superficial dermatophyte infections. Stillanother medication is NonyX, an over-the-counter keratin-dissolvingpreparation containing 9.5% ethanoic acid. It is claimed that thispreparation can penetrate the nail and selectively attack keratindebris, which is the habitat and food supply of the fungus. Like theother two, it involves a long treatment, requiring approximately 4-12months to eliminate fungus under nails, and is effective in only afraction of patients.

[0215] Because the available medications are intended for use underconditions where drug does not have ready access to the fungus, it isnot presently known what amounts and concentrations of such drugs shouldbe used with delivery through microconduits. For example, the usualpreparation containing ethanoic acid may be too strong for directapplication through nail microconduits. In the examples of Penlac® andLoprox®, both contain ciclopirox, but the need for soaking through thenail may have required that the antifungal percentage be increased by afactor of 10 over the Loprox®, in order to deliver an adequate amount tothe nail bed, to access the fungus under the nail. Since Loprox® isspecifically intended for topical use on skin, of the examplesconsidered above, perhaps it would be a good candidate for applicationthrough the nail microconduits. Tinactin®, containing the drugtolnaftate, and used for athletes foot, is also a good initial candidatefor use with nail microconduits, since it presumably isn't tooirritating in the case of direct dermal application.

[0216] Determination of how much of each drug should applied at whattime intervals through various numbers of microconduits can beinvestigated using existing methods of clinical study. The microconduitswill act additionally as drug reservoirs, such that the nail bed at thebottom of each microconduit will be exposed continuously to the drug. Ingeneral, prolonged contact with various drug preparations are desiredand acceptable. Depth and spacing of microconduits determine where thedrug will diffuse, and whether it can also be transported by a pressuredifference and electrical fields. If needed, residual drug within amicroconduit could be removed at intervals, and replaced with fresh drugcontaining solution, cream, lacquer or gel.

[0217] Microconduits can be covered with simple, protective materialssuch as plastic sheet with adhesive, nail polishes, lacquers and thelike, or left open to the air. Such covering could be removed, forexample, upon bathing, which would tend to wash out the microconduits,that could be reloaded and recovered. Alternately, microconduits cansimply be left “open.”

EXAMPLE 11 Nail 6

[0218] This experiment included forming microconduits in the toe nailsof Subject B, who had a persistent toe nail fungal infection.

[0219] Materials and Methods: A clipping of a nail on Subject B's leftmiddle toe, which was 0.024-0.025 inch thick, was made. Conditions formicroconduit formation experiments included a pressure of 25 PSI, flowsetting of 85V (85 volt) for the microparticle supply device (the custommodified S.S. White machine described earlier), and nozzle-to-nailspacing of 0.030 inch (these parameters used for the followingexperiments). It was determined that 24-26 seconds were needed toscission a microconduit through a clipping by microparticle impingement.The corresponding microconduit depth increase rate was about 0.001 inch(25u 25 micrometer) per second, which is similar to what was foundpreviously for Subject A's finger nails.

[0220] Next, the “saddle” device was attached to Subject B's LH big toenail, which had a good sized overhang (“cantilever” ); also much of thenail was detached (lifted up) from the nail bed due to the fungalcondition. The goal of this experiment was to form a microconduitthrough the overhang portion of the nail, and to determine thecorresponding scission rate. The scission rate was about one mil persecond, and the microconduit diameter was around 25 mils at top. Thisgave an initial bench mark scission rate for Subject B's nails.

[0221] Results: Subject B then selected a spot near the good-sizedoverhang on his right big toe, where the nail was well adhered to thenail bed and very near the edge of the overhang. It was decided to(microlocally) scission to a certain depth there, hopefully just gettingthrough the nail, and therefore a microconduit which reaches just to thebed (top surface of the living tissue under the nail). This nailoverhang measured 0.032-0.033 inch thick nearby, so the goal was toapproximate that time, assuming from the test scission rates that it waspossible to just reach the bed. Using the marked and calibratedhypodermic needle “yardstick” (microconduit depth probe) it wasdetermined that the hole was 800 μm (>800 micrometer) deep. This meansthat either the nail is thicker where it adhered to the bed, or itsscission rate is slower when the nail is healthy and adhered to the bed.In measuring the hole depth, Subject A pressed the slightly bluntedmicroconduit depth probe tip into the bottom. It was unyielding, feelinglike it was being pressed on nail material, unlike how it felt againstthe bottom of Subject A's “shallow” LH ring finger microconduit madeearlier, and “give” (a slight sponginess) was detected. In the presentexperiment with Subject B's toenail, however, Subject B reported nosensation even when Subject A pressed harder. All of this togetherindicated that Subject B's toe nail was clearly thicker at the“nail-attached” site.

EXAMPLE 12 Nail 7

[0222] This experiment included formation of microconduits in the toenails of Subject B, within infected regions that had thicker nail, and ademonstration of a condition for self-limiting of microconduit depth.

[0223] Materials and Methods: Prior to carrying out these experimentsmodifications were made to the nozzle holder, trying to make it easierto see the locus of the scission site by drilling view ports in thesaddle. It was determined that the set up should be as before.Microparticle impingement was carried out for a longer time, untilsensation was detected. It was decided to try some electrical resistanceexperiments. Those experiments were begun, and the following data onSubject B's RH foot, big toe, (site of the above-mentioned scission pitthat was made several days earlier) was collected. (EKG electrode onbottom of the same big toe, the 0.006 inch diameter tungsten wire as aprobe). PROBE CONDITION LOCATION RESISTANCE On toe skin saline ¼ inchbehind nail 19 kohm On nail surface dry center of nail 3.9-4 Megohm(clearly on bed) Next to saline nail appears to be 7 Megohm microconduiton bed In microconduit dry nail appears to be 7 Megohm on bed Inmicroconduit saline (within 7 Megohm microconduit)

[0224] Results: The part of the nail attached to the bed had a lowerresistance; the part of the nail that was lifted away from the bed hadvery high resistance.

[0225] The saddle/nozzle device to make a microconduit was set-up usingthe 25 psi, 85V flow setting, 0.030+ inch nozzle to nail spacing. It wasdecided to scission for 40+ seconds, or perhaps until sensation onset.The microparticle impingement was stopped at 75 seconds because ofuncertainty regarding microconduit depth. There was indeed amicroconduit, but there was evidence that the position of thesaddle/nozzle device had moved, as one could see particulate-causedhazing and a depression in front of the hole. Upon examination, thisnon-nail penetrating microconduit was found to be deeper than themicroconduit made a week ago. It was decided to form anothermicroconduit.

[0226] An attempt was made to make the saddle/nozzle steadier this time,and to tip it forward over the toe, to make certain the nozzle was aboutperpendicular to the surface of the curved nail. In this experimentmicroparticles were locally impinged for 90 seconds before terminating.Presumably, the growing microconduit had not reached living tissue.Subject A probed the bottom of the microconduit with the “depth probe”,and it felt hard. The marks on the depth probe indicated that themicroconduit was 0.040 inch (1000 micrometers) deep. A saline electricalconductivity test was performed and the resistance was in the 7 Megohmrange, indicating lots of nail left (i.e. the nail had not beenpenetrated by this microconduit).

EXAMPLE 13 Nail 9

[0227] The next experiment was for Subject B to control the particulategenerator (modified S.S. White machine) and to time the scissionduration, while Subject A held the saddle/nozzle arrangement firmly tohis toe nail, to prevent any movement. Subject B terminated theexperiment after 240 seconds. The microconduit was not through the nail.It measured a bit more than 0.040 inch (perhaps 0.045 inch) deep, washard bottomed and sensationless according to Subject B.

[0228] The particulate size, particulate nozzle diameter, microconduitdiameter are all believed to affect this self-limitation. It was decidedthat the next experiment should use the 0.018 inch nozzle instead of the0.011 inch employed in all of these experiments. This would change theform factor from 0.022 inch diameter and 0.042 inch deep (2 to 1).Extrapolation of those numbers predicts that if the nozzle is 0.018 inchdiameter, we would stall out at 0.036 inch diameter and 0.072 inch deep.Another microconduit was formed at a new site over a region attached tothe nail bed.

[0229] Using the 0.018 nozzle, with the holding fixture firmly fixed tothe nail, Subject B timed the period of scission, and could turn theparticulate flow off.

[0230] Results: At 70 seconds, Subject B slightly detected sensation. At80 seconds Subject B clearly felt mild sensation and stopped theparticulate flow. Blood was clearly issuing from the microconduit, withno sensation. Based on impingement time, the microconduit depth wasestimated as 0.060 inch.

[0231] The site was rinsed with deionized water and covered withcellophane tape. After three hours, Subject B reported sensation in thattoe, and trimmed the nail overhang, the trimming stopping the sensation.There had been observable additional blood flow. The tape was removedtwo days later, with no further observable effects.

[0232] Subject A measured Subject B's toe nail overhang right in frontof these sites. The toe nail thickness was in the range of 0.050-0.065inch. In looking under his nails, there was a striated buildup of whatappeared to be fungally-altered nail material. The middle toe on hisright foot was the worst. The altered material ranged between 0.125 and0.200 inch thick. Protruding grey/white regions of material are punky,but quite hard in compression.

EXAMPLE 14 Nail 10

[0233] This Experiment created a microconduit in the toe nail of SubjectA and demonstrates ‘nail piercing’ to accomodate ‘nail jewelry’.

[0234] Methods and Materials: The experiment was carried out on SubjectB's right hand big toe nail. A location in the overhang of the nail,near a protruding mass of fungal-infected nail material and debris,measuring 0.060-0.090 inches thick was selected.

[0235] Results: Particulate impingement for 240 seconds created amicroconduit through the nail. A piece of 0.005 inch tungsten wire wasinserted through the microconduit, around the nail edge, and backthrough the microconduit, thus inventing nail jewelry. This decorationwas duplicated with a piece of gold wire. This demonstrates ‘nailpiercing’ for cosmetic reasons.

[0236] Also, Subject B demonstrated the use of a toe nail microconduitas a reservoir for drug-containing cream (Micronazole Nitrate Cream, USP2% (E. Fugera & Co., Melville, NY) by spreading the cream over the locusof the microconduit and wiping off the excess. This left a small ‘whitedot’ that could easily be reloaded as desired.

[0237] Equivalents

[0238] While this invention has been particularly shown and describedwith references to certain embodiments thereof, it will be understood bythose skilled in the art that various changes in form and details may bemade therein without departing from the spirit and scope of theinvention encompassed by the appended claims. Such equivalents areintended to be encompassed in the scope of the following claims.

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What is claimed is:
 1. A method for forming at least one microconduit ina tissue, comprising the steps of: a) accelerating a plurality ofmicroparticles to a velocity that causes the microparticles to penetratea region of a tissue surface upon impingement of the microparticles onthe tissue surface; b) directing the microparticles towards the regionof tissue surface, thereby causing the microparticles to penetrate thetissue; and c) scissioning the tissue with the impinging microparticles,thereby forming a plurality of free microtissue particles, and therebyforming a microconduit.
 2. The method of claim 1, wherein themicroparticles are accelerated by a flowing gas.
 3. The method of claim2, wherein the flowing gas is at a pressure greater than about oneatmosphere absolute.
 4. The method of claim 2, wherein the flowing gasis at a pressure of less than about one atmosphere absolute.
 5. Themethod of claim 1, wherein the microparticles are accelerated by meansof a flowing liquid.
 6. The method of claim 5, wherein the flowingliquid is at a pressure greater than about one pound per square inch. 7.The method of claim 5, wherein the flowing liquid is at a temperature ofbelow about 20° C.
 8. The method of claim 1, wherein the microparticlesare accelerated by contacting the microparticles with a moving, solidsurface.
 9. The method of claim 8, wherein the moving solid surfaceincludes a rotating impeller.
 10. The method of claim 1, wherein themicroparticles are directed through at least one microhole in a mask atthe tissue surface, the mask comprising a membrane.
 11. The method ofclaim 10, wherein the membrane is conformable to the tissue surface. 12.The method of claim 1, wherein a plurality of microparticles areaccelerated and directed to the region of tissue surface, and whereinthe microparticles are collimated to form a beam of collimatedmicroparticles.
 13. The method of claim 12, wherein the collimated beamof microparticles is scanned over the region of tissue surface.
 14. Themethod of claim 13, wherein the collimated beam of microparticles isgated on and off.
 15. The method of claim 1, wherein the microparticlesare accelerated to a velocity sufficient to cause tissue material to bemicroscissioned from the tissue by impingement of the microparticles onthe tissue surface.
 16. The method of claim 15, wherein the tissuematerial that is microscissioned is ejected from the tissue surface. 17.The method of claim 1, wherein the microparticles are accelerated to avelocity sufficient to cause tissue material to be compacted when themicroparticles penetrate the tissue.
 18. The method of claim 1, whereinthe microparticles impinge upon the region of tissue surface which hasan area in a range of between about 100 square micrometers and about twomillion square micrometers.
 19. The method of claim 1, further includingthe step of modifying at least one physical property of the region oftissue surface.
 20. The method of claim 1, further including the step ofapplying at least one electrical pulse to cause electroporation of atleast one lipid-containing membrane of the region of tissue surface. 21.The method of claim 20, wherein the electrical pulse is appliedfollowing formation of the microconduit.
 22. The method of claim 1,further including the step of applying a direct current voltage to themicroconduit to produce iontophoresis.
 23. The method of claim 22,wherein the direct current voltage applied to the microconduit ispulsed.
 24. The method of claim 1, wherein the microparticles are notsoluble in water.
 25. The method of claim 1, wherein the microparticlesare soluble in water.
 26. The method of claim 25, wherein themicroparticles include a therapeutically effective substance.
 27. Themethod of claim 1, wherein the microparticles include aluminum oxide.28. The method of claim 1, wherein the microparticles include sodiumbicarbonate.
 29. The method of claim 1, wherein the microparticlesinclude urea.
 30. The method of claim 1, wherein the microparticlesinclude solid carbon dioxide.
 31. The method of claim 1, wherein themicroparticles include solid water.
 32. The method of claim 1, whereinthe melting point of the microparticles is less than about 33° C. 33.The method of claim 32, wherein the microparticles include at least onetherapeutically effective substance.
 34. The method of claim 2, whereinthe flowing gas includes air.
 35. The method of claim 2, wherein theflowing gas is at a temperature of below about 20° C.
 36. The method ofclaim 2, wherein the flowing gas includes an inert gas.
 37. The methodof claim 1, further including the step of applying a chemical agent tothe microconduit that affects a rate of recovery of the microconduit.38. The method of claim 37, wherein the chemical agent includes acalcium ion.
 39. The method of claim 37, wherein the chemical agentincludes 5-fluorouracil.
 40. The method of claim 37, wherein thechemical agent is selected from the group consisting of retinoids,surfactants, and antigents.
 41. The method of claim 40, wherein thechemical agent includes retinoic acid.
 42. The method of claim 37,further including the step of directly applying pressure to themicroconduit through a column containing the chemical agent, the columnsealed to the tissue around the microconduit.
 43. The method of claim 1,further including the step of testing for the presence of blood withinthe microconduit.
 44. The method of claim 43, wherein the test employsoptical means.
 45. The method of claim 44, wherein the optical meansincludes image analysis.
 46. A method for forming at least one openingin the stratum corneum of skin comprising the steps of: a) acceleratinga plurality of microparticles to a velocity that causes themicroparticles to penetrate a region of a skin surface upon impingementof the microparticles on the skin surface; b) directing themicroparticles towards the region of skin surface, thereby causing themicroparticles to penetrate the skin; and c) scissioning the skin withthe impinging microparticles, thereby forming a plurality of freemicrotissue particles, and thereby forming a microconduit in the skin.47. The method of claim 46, wherein the microparticles include at leastone therapeutically effective substance.
 48. The method of claim 46,wherein the microparticles are accelerated by a flowing gas.
 49. Themethod of claim 48, wherein the flowing gas is at a pressure greaterthan about one atmosphere absolute.
 50. The method of claim 48, whereinthe flowing gas is at a pressure of less than about one atmosphereabsolute.
 51. The method of claim 46, wherein the microparticles areaccelerated by means of a flowing liquid.
 52. The method of claim 51,wherein the flowing liquid is at a pressure greater than about one poundper square inch.
 53. The method of claim 51, wherein the flowing liquidis at a temperature of below about 20° C.
 54. The method of claim 46,wherein the microparticles are accelerated by contacting themicroparticles with a moving, solid surface.
 55. The method of claim 54,wherein the moving solid surface includes a rotating impeller.
 56. Themethod of claim 46, wherein the microparticles are directed through atleast one microhole in a solid mask at the skin surface.
 57. The methodof claim 46, wherein the microparticles that are directed to the regionof skin surface are collimated to form a beam of collimatedmicroparticles.
 58. The method of claim 57, wherein the beam ofcollimated microparticles is scanned over the region of skin surface.59. The method of claim 58, wherein the beam of collimatedmicroparticles is gated on and off.
 60. The method of claim 46, whereinthe microparticles are accelerated to a velocity sufficient to causeskin material to be microscissioned from the skin by impingement of themicroparticles on the skin surface.
 61. The method of claim 60, whereinthe skin material that is microscissioned is ejected from the skinsurface.
 62. The method of claim 46, wherein the microparticles areaccelerated to a velocity sufficient to cause skin material to becompacted when the microparticles penetrate the skin.
 63. The method ofclaim 46, wherein the microparticles impinge upon a region of skinsurface having an area equal to between about 1000 square micrometersand about 100,000 square micrometers.
 64. The method of claim 46,further including the step of modifying at least one physical propertyof the region of skin surface.
 65. The method of claim 46, furtherincluding the step of applying at least one electrical pulse to causeelectroporation of at least one lipid-containing membrane of the skin tothereby cause formation of at least one aqueous pathway.
 66. The methodof claim 65, wherein the electrical pulse is applied following formationof the microconduit.
 67. The method of claim 65, further comprising thestep of applying at least one modifying agent that alters the aqueouspathway.
 68. The method of claim 46, further including the step ofapplying a direct current voltage to the microconduit to produceiontophoresis.
 69. The method of claim 68, wherein the direct currentvoltage applied to the microconduit is pulsed.
 70. The method of claim46, wherein the microconduit fully penetrates a stratum corneum layer ofthe skin.
 71. The method of claim 70, wherein the microconduit furtherpenetrates the epidermis.
 72. The method of claim 70, wherein themicroconduit further penetrates the dermis.
 73. The method of claim 46,further comprising the step of making at least one measurement of theamount of water vapor in a gas exiting from the microconduit duringformation of the microconduit.
 74. A method of delivering a therapeuticmolecule or ion to tissue, comprising the steps of: a) accelerating aplurality of microparticles to a velocity that causes the microparticlesto penetrate a region of a tissue surface upon impingement of themicroparticles on the tissue surface; b) directing the microparticlestowards the region of tissue surface, thereby causing the microparticlesto penetrate the tissue; c) scissioning the tissue with the impingingmicroparticles, thereby forming a plurality of free microtissueparticles, and thereby forming a microconduit; and d) administering atleast one therapeutic molecule or ion by directing the therapeuticmolecule or ion into at least one microconduit, thereby delivering atherapeutic molecule or ion to tissue.
 75. The method of claim 74,comprising the further step of directly applying pressure to themicroconduit through a column containing the therapeutic molecule orion, the column sealed to the tissue around the microconduit.
 76. Themethod of claim 75, wherein the pressure that is applied is a pressuregradient.
 77. The method of claim 74, wherein the therapeutic moleculeor ion is in a controlled release material.
 78. The method of claim 74,wherein the therapeutic molecule or ion is supplied within a hydrogel,and the hydrogel is administered by directing the hydrogel into themicroconduit, thereby delivering the therapeutic molecule or ion to thetissue.
 79. The method of claim 74, wherein the therapeutic molecule orion is an immunizing material.
 80. The method of claim 74, wherein thetherapeutic molecule or ion is a nucleic acid or a modified nucleicacid.
 81. The method of claim 80, wherein the nucleic acid is DNA.
 82. Amethod of extracting an analyte from a tissue, comprising the steps of:a) accelerating a plurality of microparticles to a velocity that causesthe microparticles to penetrate a region of a tissue surface uponimpingement of the microparticles on the tissue surface; b) directingthe microparticles towards the region of tissue surface, thereby causingthe microparticles to penetrate the tissue; c) scissioning the tissuewith the impinging microparticles, thereby forming a plurality of freemicrotissue particles, and thereby forming a microconduit; and d)removing the analyte from the tissue through the microconduit, therebyextracting the analyte from the tissue.
 83. The method of claim 82,wherein the analyte is removed by sampling or by reducing pressure overthe microconduit.
 84. The method of claim 82, further comprising thestep of measuring the amount of analyte while the analyte is within themicroconduit.
 85. The method of claim 82, wherein the tissue is skin.86. The method of claim 84, wherein the tissue is skin.
 87. A method forforming a molecular matrix within at least one microconduit, comprisingthe steps of: a) accelerating a plurality of microparticles to avelocity that causes the microparticles to penetrate a region of atissue surface upon impingement of the microparticles on the tissuesurface; b) directing the microparticles towards the region of tissuesurface, thereby causing the microparticles to penetrate the tissue; c)scissioning the tissue with the impinging microparticles, therebyforming a plurality of free microtissue particles, and thereby forming amicroconduit; and c) directing a molecular matrix into the microconduit,thereby forming a molecular matrix within the microconduit.
 88. Themethod of claim 87, wherein the molecular matrix is a gel.
 89. Themethod of claim 88, wherein the gel is calcium alginate.
 90. The methodof claim 88, wherein the molecular matrix is a polymer matrix.
 91. Amethod of transdermal delivery of a therapeutic molecule or ion,comprising the steps of: a) accelerating a plurality of non-drugcontaining microparticles to a velocity that causes the microparticlesto completely penetrate a region of a skin surface upon impingement ofthe microparticles on the skin surface; b) directing the microparticlestowards the region of the skin surface, thereby causing themicroparticles to penetrate the skin; c) scissioning the skin with theimpinging microparticles, thereby forming a plurality of freemicrotissue particles, and thereby forming a microconduit; and d)administering at least one therapeutic molecule or ion by directing thetherapeutic molecule or ion into at least one microconduit, therebydelivering therapeutic molecule or ion through the stratum corneum. 92.The method of claim 91, comprising the further step of directly applyingpressure to the microconduit through a column containing the therapeuticmolecule or ion, the column sealed to the tissue around themicroconduit.
 93. The method of claim 91, wherein the therapeuticmolecule or ion is in a controlled release material.
 94. The method ofclaim 91, wherein the therapeutic molecule or ion is supplied within ahydrogel, and the hydrogel is administered by directing the hydrogelinto at least one microconduit, thereby delivering a therapeuticmolecule or ion through the stratum corneum.
 95. The method of claim 91,further including the step of applying an electric field oriented in adirection to cause the therapeutic molecule or ion to migrate from themicroconduit into the skin and parallel to a major plane of the regionof skin surface.
 96. The method of claim 91, further including the stepof applying a stimulus to the skin that causes uptake of the therapeuticmolecule or ion into at least one cell within the skin.
 97. The methodof claim 96, wherein the cell is a dendritic cell.
 98. The method ofclaim 96, wherein the therapeutic molecule or ion is DNA or ananti-neoplastic drug.
 99. The method of claim 46, further including thestep of making at least one measurement of electrical impedance of theregion of skin surface to monitor formation of the microconduit duringimpingement by the microparticles.
 100. A method for making at least onebiopotential measurement across the skin, comprising the steps of: a)accelerating a plurality of microparticles to a velocity that causes themicroparticles to penetrate a region of a skin surface upon impingementof the microparticles on the skin surface; b) directing themicroparticles towards the region of skin surface, thereby causing themicroparticles to penetrate the skin; c) scissioning the skin with theimpinging microparticles, thereby forming a plurality of freemicrotissue particles, and thereby forming a microconduit; d) placing atleast two electrodes in electrical connection with the skin with atleast one electrode at the microconduit; and e) making a biopotentialmeasurement across the skin.
 101. The method of claim 100, wherein thebiopotential measurement is an electrocardiogram.
 102. The method ofclaim 101, wherein the electrocardiogram measurement is obtained duringexercise stress testing.
 103. The method of claim 100, wherein thebiopotential measurement is an electromyogram.
 104. The method of claim100, wherein the biopotential measurement made is suitable forneuromuscular testing.
 105. The method of claim 100, wherein thebiopotential measurement is an electroencephalogram to monitoranaesthesia.
 106. A method of delivering at least one molecule to tissuecomprising the step of storing the molecule in at least one puncturablecapsule in proximity to at least one microconduit.
 107. The method ofclaim 106, wherein the stored molecule includes a therapeutic moleculeor ion.
 108. A mask for defining at least one localized area of a tissuesurface region for formation of a microconduit by microparticleimpingement, the mask comprising: a) a membrane that has a thickness ina range of between about one micrometer and about one thousandmicrometers; b) at least one microhole in said membrane, the microholehaving a diameter in a range of between about three micrometers andabout one thousand micrometers; and c) means for positioning saidmembrane on a tissue surface.
 109. The mask of claim 108, wherein themembrane is conformable to the tissue surface.
 110. A method for formingat least one microconduit through nail tissue, comprising the steps of:a) accelerating a plurality of microparticles to a velocity that causesthe microparticles to penetrate a region of nail tissue surface uponimpingement of the microparticles on the nail tissue surface; b)directing the microparticles towards the region of nail tissue surface,thereby causing the microparticles to penetrate the nail tissue surface;and c) scissioning the nail tissue with the impinging microparticles,thereby forming a plurality of free nail microtissue particles, andthereby forming a microconduit through the nail tissue.
 111. A methodfor treating an infection of tissue underlying nail tissue, comprisingthe steps of: a) accelerating a plurality of microparticles to avelocity that causes the microparticles to penetrate a region of nailtissue surface upon impingement of the microparticle on the nail tissuesurface; b) directing the microparticles towards the region of nailtissue surface, thereby causing the microparticles to penetrate the nailtissue surface; c) scissioning the nail tissue with the impingingmicroparticles, thereby forming a plurality of free nail microtissueparticles, and thereby forming a microconduit through the nail tissue;and d) administering at least one therapeutic molecule or ion bydirecting the therapeutic molecule or ion into at least onemicroconduit, thereby delivering the therapeutic molecule or ion throughthe nail tissue.
 112. A method for marking nail tissue with at least oneidentifying mark or at least one decorative mark, comprising the stepsof: a) accelerating a plurality of microparticles to a velocity thatcauses the microparticles to partially penetrate into a region of nailtissue surface upon impingement of the microparticles on the nail tissuesurface; b) directing the microparticles towards the region of nailtissue surface, thereby causing the microparticles to partiallypenetrate the nail tissue surface and form a microconduit in the nailtissue; and c) scissioning the nail tissue with the impingingmicroparticles, thereby forming a plurality of free nail microtissueparticles, and thereby forming a microconduit through the nail tissue;and d) directing a dye or an ink into at least one microconduit, therebymarking the nail tissue.
 113. A method for inserting at least one wirethrough at least one microconduit, comprising the steps of: a)accelerating a plurality of microparticles to a velocity that causes themicroparticles to penetrate a region of nail tissue surface uponimpingement of the microparticles on the nail tissue surface; b)directing the microparticles towards the region of nail tissue surfacethat extends beyond the body, thereby causing the microparticles topenetrate the nail tissue surface; c) scissioning the nail tissue withthe impinging microparticles, thereby forming a plurality of free nailmicrotissue particles, and thereby forming a microconduit through thenail tissue; and d) directing a wire into at least one microconduit,thereby inserting the wire through the microconduit.
 114. A method ofreducing a pressure caused by a pool of blood beneath an injured ortraumatized nail comprising the steps of: a) accelerating a plurality ofmicroparticles to a velocity that causes the microparticles to penetratea region of nail tissue surface upon impingement of the microparticleson the nail tissue surface; b) directing the microparticles towards theregion of nail tissue surface, thereby causing the microparticles topenetrate the nail tissue surface; c) scissioning the nail tissue withthe impinging microparticles, thereby forming a plurality of free nailmicrotissue particles, and thereby forming a microconduit through thenail tissue; and d) thereby releasing the pressure through themicroconduit.